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United States Patent Application 20110245592
Kind Code A1
Schoolcraft; William B. ;   et al. October 6, 2011

Secretome Profile-Facilitated In Vitro Fertilization

Abstract

Secretome profiling improves the pregnancy success rate of in vitro fertilization processes, while reducing the risk of multiple births.


Inventors: Schoolcraft; William B.; (Englewood, CO) ; Katz-Jaffe; Mandy; (Denver, CO) ; McReynolds; Susanna; (Parker, CO)
Serial No.: 081463
Series Code: 13
Filed: April 6, 2011

Current U.S. Class: 600/34; 435/287.1; 435/287.2
Class at Publication: 600/34; 435/287.1; 435/287.2
International Class: A61B 17/435 20060101 A61B017/435; C12M 1/34 20060101 C12M001/34


Claims



1. A system for enhancing the pregnancy success rate of in vitro fertilization, comprising: means for determining the secretome profile of an embryo to identify proteins implicated in implantation success; and means for recommending whether to implant the embryo on the basis of the secretome profile.

2. The system of claim 1, further comprising data representative of a secretome profile of an embryo acquired by ELISA.

3. The system of claim 2, further comprising the secretome profile of an embryo acquired using a secretome panel selected from the group consisting of SEQ ID Nos. 1-403.

4. The system of claim 3, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 1-404.

5. The system of claim 3, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 261-404.

6. The system of claim 3, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 317, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 383, 391, 397, 398, 399, 402, 403, and 404.

7. The system of claim 3, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 391, 397, 398, 399, 402, 403, and 404.

8. The system of claim 3, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 317 and 383.

9. The system of claim 1, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 1-404.

10. The system of claim 1, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 261-404.

11. The system of claim 1, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 317, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 383, 391, 397, 398, 399, 402, 403, and 404.

12. The system of claim 1, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 391, 397, 398, 399, 402, 403, and 404.

13. The system of claim 1, further comprising the secretome profile of an embryo acquired using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 317 and 383.

14. A method of in vitro fertilization, comprising: determining the secretome profile of a candidate embryo to ascertain proteins implicated in implantation success; modeling on basis of the secretome profile to assess candidate embryo viability for recommending whether to implant the candidate embryo on the basis of the viability assessment; and conditionally implanting the candidate embryo on the basis of the recommendation.

15. The method of claim 14, wherein the step of determining the secretome profile includes using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 1-404.

16. The method of claim 14, wherein the step of determining the secretome profile includes using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 261-404.

17. The method of claim 14, wherein the step of determining the secretome profile includes using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 317, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 383, 391, 397, 398, 399, 402, 403, and 404.

18. The method of claim 14, wherein the step of determining the secretome profile includes using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 310, 311, 313, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 391, 397, 398, 399, 402, 403, and 404.

19. The method of claim 14, wherein the step of determining the secretome profile includes using a secretome panel selected as one or more members of the group consisting of SEQ ID Nos. 317 and 383.

20. In an ELISA assay kit comprising a plurality of microwells for the quantitation of protein content in a sample, the improvement comprising: the microwells being constructed and arranged to quantitate for a plurality of proteins selected from SEQ ID Nos. 1-404.

21. The kit claim 20, wherein the plurality of proteins are selected from the group consisting of SEQ ID Nos. 261-404.

22. The kit claim 20, wherein the plurality of proteins are selected from the group consisting of SEQ ID Nos. 310, 311, 313, 317, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 383, 391, 397, 398, 399, 402, 403, and 404.

23. The kit claim 20, wherein the plurality of proteins are selected from the group consisting of SEQ ID Nos. 310, 311, 313, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 391, 397, 398, 399, 402, 403, and 404.

24. The kit claim 20, wherein the plurality of proteins are selected from the group consisting of SEQ ID Nos. 317 and 383.
Description



RELATED APPLICATIONS

[0001] This application claims benefit of priority to U.S. Provisional Patent Application No. 61/321,448 filed Apr. 6, 2010, which is incorporated by reference to the same extent as though fully replicated herein.

SEQUENCE LISTING

[0002] This application includes a Sequence Listing, as set forth in an ASCII-compliant text file named "CCRMProtein_ST25.txt", created on Apr. 6, 2011, and containing 2263 kilobytes, which is incorporated by reference to the same extent as though fully replicated herein.

BACKGROUND

[0003] 1. Field of the Invention

[0004] The invention relates to the field of in vitro fertilization (IVF), which is a process by which mammalian egg cells are fertilized by sperm outside the womb. More particularly, evaluation of a secretome profile is used to enhance the pregnancy success rate when the fertilized egg is implanted into a patient's uterus.

[0005] 2. Description of the Related Art

[0006] IVF infertility treatment offers infertile couples a chance to have a biologically related child. IVF may overcome female infertility due to problems of the fallopian tube or endometriosis. IVF also overcomes male infertility due to problems with sperm quality or quantity. The IVF process involves hormonally controlling the ovulatory process, removing eggs (termed ova) from the woman's ovaries and permitting the sperm to fertilize the eggs in a fluid medium. The fertilized egg, termed an embryo, is subsequently transferred to the patient's uterus with the intent of establishing a successful pregnancy. Due to expensive procedural costs, IVF is only attempted after the failure of less expensive fertility treatments.

[0007] FIG. 1 displays a schematic for mammalian blastocyst implantation. Mammalian embryo implantation is a complex and intricate process involving numerous biological changes at both the embryo and endometrial level. The interaction between the blastocyst and the endometrium is a function of both a receptive endometrial environment and a healthy blastocyst. A blastocyst failing to implant or an endometrium failing to sustain growth and differentiation will result in spontaneous abortion. The prior art teaches very little about the embryo's role in the events leading to the attachment of a viable blastocyst to a receptive uterine luminal epithelium.

[0008] IVF treatment begins with administration of hormonal medications to stimulate ovarian follicle production, such as gonadotropins hormones. The prevention of spontaneous ovulation involves using other hormones, such as GnRH antagonists or GnRH agonists that block the natural surge of luteinizing hormone. With adequate follicular maturation, administration of human chorionic gonadotropin hormone causes ovulation approximately 42 hours after the administration. However, the egg retrieval procedure takes place just prior to ovulation, in order to recover the eggs from the ovary. The egg retrieval proceeds using a transvaginal technique involving an ultrasound-guided needle that pierces the vaginal wall to reach the ovaries. After recovery of the follicles through the needle, the follicular fluid is provided to the IVF laboratory to identify eggs. Typically, the procedure retrieves between 10 and 30 eggs. The retrieval procedure takes approximately 20 minutes and is usually done under conscious sedation or general anesthesia.

[0009] For IVF, the fertilization of the egg (termed insemination) proceeds in the laboratory where the identified eggs and semen are usually incubated together in a culture media. The confirmation of fertilization proceeds by monitoring the eggs for cell division. For instance, a fertilized egg may show two pronuclei. In certain situations, such as low sperm count or motility, a single sperm may be injected directly into the egg using a method called intracytoplasmic sperm injection (ICSI). In another option known as gamete intrafallopian transfer, eggs are removed from the woman and placed in one of the fallopian tubes, along with the man's sperm. In this example, fertilization occurs within the women's body, a process termed in vivo fertilization.

[0010] Selected embryos are transferred to the patient's uterus through a thin, plastic catheter, which goes through the vagina and cervix. Typically, transfer of 6-8 cell stage embryos to the uterus occurs three days after embryo retrieval. Alternatively, embryos can be placed into an extended culture system with a transfer done at the blastocyst stage at approximately five days post-retrieval. Blastocyst stage transfers often result in higher pregnancy rates. Additionally, embryonic cryopreservation, or the storage of embryos in a frozen state, is feasible until uterine transfer. For example, the first term pregnancy derived from a frozen human embryo was reported in 1984.

[0011] Despite progressively improving IVF pregnancy rates, the majority of transferred human embryos result in implantation failure. For example, Canadian clinics reported an average pregnancy rate of 35% for one cycle, but a live birth rate of only 27% in 2006. Moreover, implantation success rates may decrease with the increasing maternal age, if donor eggs are not used. Various factors are associated with implantation failure, including embryo chromosome aneuploidies related to advanced maternal age and maternal factors such as endometrium response failure to hormone regulation.

[0012] To overcome low implantation success rate, multiple embryos are commonly transferred during a single IVF procedure. The process for selecting embryos for transfer often involves grading methods developed in individual laboratories to judge oocyte and embryo quality. An arbitrary embryo score, involving the number and quality of embryos, may reveal the probability of pregnancy success post-transfer. For example, an embryologist may grade embryos using morphological qualities including the number of cells, clearness of cytoplasm, evenness of growth and degree of fragmentation. However, embryo selection based on morphological qualities is not precise. Often, several embryos selected for these general qualities are implanted to improve the chance of pregnancy. The number of embryos transferred depends upon the number available, the age of the woman and other health and diagnostic factors.

[0013] The transfer of multiple embryos, however, often results in multiple pregnancies, a major complication of IVF. In general, multiple pregnancies, specifically, more than twins, hold maternal and fetal risks. For example, multiple births are associated with increased risk of pregnancy loss, neonatal morbidity, obstetrical complications, and prematurity with potential for long term damage. Some countries implemented strict limits on the number of transferred embryos to reduce the risk of high-order multiples (e.g., triplets or more). However, these limitations are not universally followed or accepted.

SUMMARY

[0014] In one embodiment, a system for enhancing the pregnancy success rate of in vitro fertilization includes a means for determining the secretome profile of an embryo to identify proteins, polypeptides, oligopeptides or protein fragments implicated in implantation success and a means for recommending whether to implant the embryo, such as a blastocyst, on the basis of the secretome profile.

[0015] The CDC data from 2006 shows that use of donor eggs in a 40 year old woman will result in a live birth 54% of the time. This compares to 20.6% using the woman's own eggs. Thus, an opportunity exists for the instrumentalities of this disclosure to facilitate success rates approximating the use of donor eggs when using one's own eggs, and may even permit higher success rates due to better selection of viable embryos.

[0016] In an embodiment, a system is provided for enhancing the pregnancy success rate of in vitro fertilization. The system includes an electronic system to configured to gather secretome data as a secretome profile of an embryo by quantitating proteins implicated in implantation success. A model is provided for use in recommending whether to implant the embryo on the basis of this secretome profile. The secretome date may be, for example, provided by use of mass spectroscopy or ELISA measurements.

[0017] According to one aspect of the system, the secretome profile may be provided by using proteins as embryo secretions in culture media that may be linked to changed odds of implantation success, for example, as may be found in one or more sequences found in SEQ ID Nos. 1-404. The sequences of SEQ ID Nos. 261-404 are preferred for this use. Particularly preferred are the sequences of SEQ ID Nos. 310, 311, 313, 317, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 383, 391, 397, 398, 399, 402, 403, and 404. The sequences of SEQ ID Nos. 310, 311, 313, 318, 319, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 345, 346, 347, 348, 349, 350, 368, 371, 374, 391, 397, 398, 399, 402, 403, and 404 are associated with increased odds of implantation success, whereas those of SEQ ID Nos. 317 and 383 are associated with failure or aneuploidy.

[0018] In an embodiment, a method of in vitro fertilization entails determining the secretome profile of a candidate embryo by use of these sequences to ascertain proteins implicated in implantation success. This provides data that may be submitted to a model that associates one or more of these proteins with changes odds of implantation success or failure. A recommendation for implantation of the embryo may then be provided based upon the modeling outcome. The embryo may be conditionally implanted on the basis of the recommendation.

[0019] In an embodiment, there is an improved ELISA assay kit with a plurality of microwells for the quantitation of protein content in a sample. The microwells are constructed and arranged to quantitate for a plurality of proteins selected from these sequences.

BRIEF DESCRIPTION OF THE FIGURES

[0020] FIG. 1 is a diagram illustrating the process blastocyst implantation.

[0021] FIG. 2 is a table that illustrates the success rates from donor oocyte IVF cycles.

[0022] FIG. 3 is a flow chart for a method of developing and using a predictive model that may be used to assess embryo viability as a candidate for use in an IVF implantation procedure.

[0023] FIG. 4 shows a system that analyzes the secretome of an embryo to assess viability.

DETAILED DESCRIPTION

[0024] The following definitions are provided to facilitate understanding of certain terms used herein and are not meant to limit the scope of the present disclosure.

[0025] The term "secretome panel" refers to a collection of individual proteins, polypeptides, oligopeptides or protein fragments that are differentially expressed and secreted by an embryo. The proteins, polypeptides, oligopeptides or protein fragments of the secretome panel are selected based on predictions of developmental competence and implantation potential of an embryo.

[0026] The term "spent media" refers to media surrounding an embryo that accumulates proteins secreted from the embryo.

[0027] The term "secretome profiling" refers to qualitative and quantitative analysis of a secretome panel collected from spent media.

[0028] There will now be shown and described a system for enhancing the pregnancy success rate of in vitro fertilization involving non-invasive secretome profiling of an embryo. Secretome profiling may be performed on embryos at differing embryonic developmental stages, such as day one through day six embryos.

[0029] Non-limiting examples of proteins comprising the secretome panel include proteins involved in amino acid metabolism, lipid metabolism, carbohydrate metabolism, signal transduction, apoptosis, transcription and combinations thereof. In one example, an embryo secretes a protein that is involved in amino acid metabolism, thereby indicating potential for successful embryo implantation. In another example, an embryo secretes a protein that is involved in apoptosis, thereby indicating a reduced potential for successful blastocyst implantation. Nonlimiting examples of the number of proteins comprising the secretome panel includes .gtoreq.250 proteins, .gtoreq.100 proteins, .gtoreq.50 proteins, .gtoreq.20 proteins or .gtoreq.10 proteins.

[0030] Secretome profiling involves assessment of a secretome panel. Secretome profiling is a noninvasive method for predicting developmental competence and viability of an embryo. Comparison of secretome profiles between developing embryos and degenerating embryos, both at the same developmental stage, reveals significant differences in protein expression. Moreover, secretome profiling provides a molecular perspective of the functioning biochemical pathways present during embryo development. A noninvasive secretome profiling assay correlates embryonic secretome to embryonic viability, thereby facilitating single embryo transfer during in vitro fertilization.

[0031] FIG. 2 highlights the importance of embryo developmental competence by showing high success rates from donor oocyte IVF cycles using young reproductive age oocytes transferred to an advanced maternal age endometrium. The data is provided for a Colorado clinic from 2004-2009 including outcomes for over 1000 donor oocyte cycles. These results demonstrate a 66.6% implantation rate for IVF recipients with an average endometrial age of 40.6 years and donor oocytes with an average age of 26.6 years.

[0032] In one embodiment, secretome profiling occurs by assessing a secretome panel via Enzyme-Linked Immunosorbent Assay (ELISA). In another embodiment, secretome profiling proceeds by assessing a secretome panel via mass spectrometry.

[0033] Non-limiting examples of solid-state substrates used in ELISA include microwell plates, such as 96-well plates, 384-well plates and 8-well strips, microarray slides and nitrocellulose membranes. In one embodiment, glass microarray surfaces contain chemical functional groups such as epoxy, amine or aldehyde. In one embodiment, microwell plate material comprises polystyrene.

[0034] The term "capture antibody" refers to an antibody secured, either covalently or non-covalently, to a solid-state substrate. The capture antibody recognizes and binds to a specific antigen such as a protein, a polypeptide, an oligopeptide, a protein fragment, a carbohydrate or a small molecule. In one example, a solid-state substrate is functionalized with capture antibodies by passive adsorption or by specific binding. For example, specific binding of capture antibodies may occur using biotinylated capture antibodies and streptavidin-coated solid-state substrates. In another example, solid-state substrates are functionalized with antigens via passive adsorption.

[0035] Non-limiting examples of ELISA types include direct, indirect, competitive, and sandwich. In one example, proteins from the secretome panel are detected on a solid-state substrate using either a primary labeled antibody or a secondary labeled antibody. In another example, proteins from the secretome panels are detecting using a sandwich ELISA wherein a protein from the secretome panel is bound between two primary antibodies, namely the capture antibody and the detection antibody. Non-limiting examples of antibody labels include enzymes and fluororphores. In one embodiment, an enzyme that is conjugated to a detection antibody binds to a substrate producing either a chromogenic, fluorescent or chemiluminescent signal that is proportional to the quantity of protein from the secretome panel.

[0036] FIG. 3 shows a method 300 implementing the concepts described above to identify those embryos having the greatest chanced for implantation success rates. Step 302 entails ascertaining the secretome of proteins secreted into culture medium by an egg, blastocyst or developing embryo (hereinafter "embryo") that has been previously fertilized by methods known to the art. Step 302 may be performed at any time, including before an egg is fertilized or immediately thereafter. More typical times to sample the media include form one to five days after fertilization occurs. Step 302 is preferably conducted before step 304, but samples of the culture medium may be frozen for later analysis. The embryo is then used 304 in an IVF procedure attempting to implant the embryo in a woman who desires to have a baby. The implantation successes and failures are monitored 306 to collect data relating the successes and failures to one or more proteins of the secretome. Once a sufficient number of instances have transpired 308 to provide for statistical significance, the proteins are mapped 310 to the instances of success and failure. The exact manner of mapping is not of particular importance, as there a number of biostatistical analysis software products are commercially available. For example, this may be done using the JMP.RTM. 9 packages from SAS of Cary, N.C. to implement methods as taught by Zar, Biostatistical Analysis, 5th Ed., Prentice Hall (2007) to map the secretome profiles using also time as a variable. The mapping may be multivariate, or by an artificially intelligent algorithm, such as a neural network. This model may be then used 312 as a predictive tool to select embryos for implantation based upon their secretome profile. This is done by analyzing 314 the secretome of candidate embryos for implantation with use of the model to predict implantation success or failure. One or more of the selected embryos are then implanted 316

[0037] FIG. 4 shows an analytical system 400 that may be used in step 314 according to one embodiment. A specimen 402 contains a sample of culture medium that is collected for secretome analysis as described above. By way of example, this specimen may contain culture medium in the form of a single or multi-well ELISA assay, or in a form that has bee pre-processed for mass spectroscopy analysis. A corresponding measurement device 404 obtains primary measurements from the specimen 402. The measurements are representative of one or more proteins in the secretome that have been mapped to incidences of implantation success of failure. The model for this mapping is provided in step 312 of method 300 and resides on analyzer 406, which may be a computer operating on program instructions. The analyzer 406 receives signals from the measurement device 404 and interprets these by computational analysis according to the model then provides the results of computational analysis to an output device 408, which may be printer or an optical display. The various components of system 400 may be partially integrated, such as by combining the measurement device 404, analyzer 406, and output device 408 into one integrated system.

[0038] It will be appreciated that perceptive use of the instrumentalities described herein may result in a better selection of healthy embryos, such as blastocysts, for implantation. Thus, fewer blastocysts need to be implemented, such that there is lower risk of multiple pregnancies while achieving a higher overall pregnancy success rate.

[0039] The following descriptions will show and describe, by way of non-limiting examples, a process for improving pregnancy success rates with lower incidence of multiple births. The following examples describe secretome profiling of spent media from human blastocysts to provide implantation recommendation. It is to be understood that these examples are provided by way of illustration and should not be unduly construed to limit the scope of what is disclosed herein.

Example 1

Secretome Profiling of Human Embryos Using Mass Spectrometry

[0040] The following nonlimiting example teaches by way of illustration, not by limitation, secretome profiling of a human embryo using mass spectrometry (MS). Human cleavage-stage embryos were cultured in 10 .mu.L drops of G1 supplemented with 2.5 mg/mL recombinant albumin under oil at 37.degree. C., 6% CO.sub.2, 5% O.sub.2 for 24 hours. The embryos were washed twice in G2 culture media and further cultured in 10 .mu.L drops of G2 supplemented with 2.5 mg/mL recombinant albumin under oil at 37.degree. C., 6% CO.sub.2, 5% O.sub.2 for 48 hours with a fresh drop of G2 media added after 24 hours. Spent media samples of blastocysts were transferred into 0.65 mL Eppendorf tubes. Control groups comprised media cultured and collected under the same conditions but without embryos.

[0041] Micro-drops of spent media were depleted of human serum albumin (HSA) using Cibracron Blue Activated SwellGel Discs (Themo Fisher Scientific, Rockford, Ill.). The proteins in the spent media were separated by 1D gel electrophoresis (Invitrogen, Carlsbad, Calif.), followed by Coomassie staining. Twenty five individual bands were cut out from each sample lane and a standard in-gel digestion protocol was used based on previously used methods ((1.) Rosenfeld, J., Capdevielle, J., Guillemot, J. C. & Ferrara, P. In-gel digestion of proteins for internal sequence analysis after one- or two-dimensional gel electrophoresis. Anal. Biochem. 203, 173-179 (1992), (2.) Hellman, U., Wernstedt, C., Gonez, J. & Heldin, C. H. Improvement of an "In-Gel" digestion procedure for the micropreparation of internal protein fragments for amino acid sequencing. Anal. Biochem. 224, 451-455 (1995)). Iodoacteamide (IAM) was used for cysteine alkylation. The samples were analyzed on a LTQ-FT Ultra Hybrid Mass Spectrometer (Thermo/Finnigan; Waltham, Mass.) with a method based on a previously described protocol from Hansen et al., (Hansen KC, Kiemele L, Maller O, O'Brien J, Shankar A, Formetti J, Schedin P. An in-solution ultrasonication-assisted digestion method for improved extracellular matrix proteome coverage. Mol Cell Proteomics. 8(7):1648-57 (2009)).

[0042] Spent media samples were analyzed on a LTQ-FT Ultra Hybrid Mass Spectrometer (Thermo/Finnigan; Waltham, Mass.). Peptide desalting and separation was achieved using a dual capillary/nano pump HPLC system (Agilent 1200, Palo Alto, Calif.). On this system 8 .mu.L of spent media sample was loaded onto a trapping column (ZORBAX 300SB-C18, dimensions 5.times.0.3 mm 5 .mu.m) and washed with 5% acetonitrile (ACN), 0.1% formic acid (FA) at a flow rate of 15 .mu.L/min for 5 minutes. At this time, the trapping column was put online with the nano-pump at a flow rate of 350 nL/min. An 85 minute gradient, from 8% ACN to 40% ACN, was used to separate the peptides. The column was made from an in-house pulled 360/100 nm (outer/inner diameter) fused silica capillary packed with Jupiter C18 resin (Penomenex; Torrance, Calif.). The column was kept at a constant 40.degree. C. using an in-house built column heater. Data acquisition was performed using the instrument supplied Xcalibur (version 2.0.6) software. The LC runs were monitored in positive ion mode by sequentially recording survey MS scans (m/z 400-2000), in the ICR cell, while three MS.sup.2 were obtained in the ion trap via CID for the most intense ions. After two acquisitions of a given ion within 45 seconds, the ion was excluded for 150 seconds.

[0043] For data analysis, The Raw Distiller program (UCSF) was used to create a de-isotoped centroided peak lists from the raw spectra into the mascot format using the default settings. The peak lists were searched against the SwissProt Human database (51.6, Homo sapiens 15720 sequences) using Mascot.TM. server (Version 2.2, Matrix Science, Boston, Mass.). The search parameters are the same as those followed in Hansen et al., (Hansen K C, Kiemele L, Maller O, O'Brien J, Shankar A, Formetti J, Schedin P. An in-solution ultrasonication-assisted digestion method for improved extracellular matrix proteome coverage. Mol Cell Proteomic. 8(7): 1648-57 (2009)). The Mascot results were loaded into Scaffold (v 2.06) and the runs were compared.

[0044] In one embodiment, MS analysis of spent media reveals a secretome panel of 261 individual proteins (Table 1). In one example, secretome profiling using the secretome panel of 261 proteins (Table 1), as analyzed by MS, identifies the potential for developmental competence and implantation success of a human embryo.

[0045] In one embodiment, analysis of spent media via MS reveals a secretome panel of 37 individual proteins (Table 2). Secretome profiling via MS of 37 individual proteins (Table 2) from spent media of human embryos correlates with embryonic viability and euploidy. The term "euploidy" refers to having a chromosome number that is an exact multiple of the haploid number for a human embryo, namely 23 pairs of chromosomes.

[0046] In Tables 1 and 2 below, the entries for "Entry Name" and "Accession Number" refer to identifiers for published sequence data that is stored in bioinformatic databases including the Uniprot Knowledgebase, Swiss-Prot and TrEMBL. The information is made freely available to the world and is coordinated by the Swiss Institute of Bioinformatics, which is centrally administered in Lausanne, Switzerland with offices in Bern, Geneva and Zurich. The sequence information represented by these identifiers together with the representative publications is hereby incorporated by reference to the same extent as though fully replicated herein. For those proteins having isoforms, the sequences for Table 1 include consensus sequences for the primary isoforms, while sequences for Table 2 include also all other isoforms available at the time of filing.

TABLE-US-00001 TABLE 1 Proteins identified in spent media of human embryos. Protein SEQ ID Molecular Accession NO. Protein Name Entry Name Weight Number Taxonomy 1 Serum albumin ALBU_HUMAN 69 kDa P02768 Homo sapiens 2 Serotransferrin TRFE_HUMAN 77 kDa P02787 Homo sapiens 3 Haptoglobin HPT_HUMAN 45 kDa P00738 Homo sapiens 4 Alpha-1-antitrypsin A1AT_HUMAN 47 kDa P01009 Homo sapiens 5 Alpha-2-macroglobulin A2MG_HUMAN 163 kDa P01023 Homo sapiens 6 Hemopexin HEMO_HUMAN 52 kDa P02790 Homo sapiens 7 Apolipoprotein A-I APOA1_HUMAN 31 kDa P02647 Homo sapiens 8 Ceruloplasmin CERU_HUMAN 122 kDa P00450 Homo sapiens 9 Vitamin D-binding protein VTDB_HUMAN 53 kDa P02774 Homo sapiens 10 Alpha-1-antichymotrypsin AACT_HUMAN 48 kDa P01011 Homo sapiens 11 Plasma protease C1 inhibitor IC1_HUMAN 55 kDa P05155 Homo sapiens 12 Alpha-1B-glycoprotein A1BG_HUMAN 54 kDa P04217 Homo sapiens 13 Hemoglobin subunit beta HBB_HUMAN 16 kDa P68871 Homo sapiens 14 Antithrombin-III ANT3_HUMAN 53 kDa P01008 Homo sapiens 15 Afamin AFAM_HUMAN 69 kDa P43652 Homo sapiens 16 Angiotensinogen ANGT_HUMAN 53 kDa P01019 Homo sapiens 17 Transthyretin TTHY_HUMAN 16 kDa P02766 Homo sapiens 18 Inter-alpha-trypsin inhibitor heavy ITIH4_HUMAN 103 kDa Q14624 Homo sapiens chain H4 19 Complement factor B CFAB_HUMAN 86 kDa P00751 Homo sapiens 20 Hemoglobin subunit alpha HBA_HUMAN 15 kDa P69905 Homo sapiens 21 Ig alpha-1 chain C region IGHA1_HUMAN 38 kDa P01876 Homo sapiens 22 Ig gamma-1 chain C region IGHG1_HUMAN 36 kDa P01857 Homo sapiens 23 Heparin cofactor 2 HEP2_HUMAN 57 kDa P05546 Homo sapiens 24 Ig gamma-2 chain C region IGHG2_HUMAN 36 kDa P01859 Homo sapiens 25 Ig kappa chain C region KAC_HUMAN 12 kDa P01834 Homo sapiens 26 N-acetylmuramoyl-L-alanine PGRP2_HUMAN 62 kDa Q96PD5 Homo sapiens amidase 27 Alpha-2-HS-glycoprotein FETUA_HUMAN 39 kDa P02765 Homo sapiens 28 Kininogen-1 KNG1_HUMAN 72 kDa P01042 Homo sapiens 29 Zinc-alpha-2-glycoprotein ZA2G_HUMAN 34 kDa P25311 Homo sapiens 30 Thyroxine-binding globulin THBG_HUMAN 46 kDa P05543 Homo sapiens 31 Corticosteroid-binding globulin CBG_HUMAN 45 kDa P08185 Homo sapiens 32 Attractin ATRN_HUMAN 159 kDa O75882 Homo sapiens 33 Pregnancy zone protein PZP_HUMAN 164 kDa P20742 Homo sapiens 34 Alpha-2-antiplasmin A2AP_HUMAN 55 kDa P08697 Homo sapiens 35 Vitronectin VTNC_HUMAN 54 kDa P04004 Homo sapiens 36 Ig lambda chain C regions LAC_HUMAN 11 kDa P01842 Homo sapiens 37 Apolipoprotein A-II APOA2_HUMAN 11 kDa P02652 Homo sapiens 38 Apolipoprotein C-III APOC3_HUMAN 11 kDa P02656 Homo sapiens 39 Insulin-like growth factor-binding ALS_HUMAN 66 kDa P35858 Homo sapiens protein complex acid labile chain 40 Complement C3 CO3_HUMAN 187 kDa P01024 Homo sapiens 41 Complement C4-B CO4B_HUMAN 193 kDa P0C0L5 Homo sapiens 42 Serum paraoxonase/arylesterase 1 PON1_HUMAN 40 kDa P27169 Homo sapiens 43 Plasma retinol-binding protein RETBP_HUMAN 23 kDa P02753 Homo sapiens 44 Carboxypeptidase N subunit 2 CPN2_HUMAN 61 kDa P22792 Homo sapiens 45 Beta-2-glycoprotein 1 APOH_HUMAN 38 kDa P02749 Homo sapiens 46 Inter-alpha-trypsin inhibitor heavy ITIH2_HUMAN 106 kDa P19823 Homo sapiens chain H2 47 Clusterin CLUS_HUMAN 52 kDa P10909 Homo sapiens 48 Protein AMBP AMBP_HUMAN 39 kDa P02760 Homo sapiens 49 Ig heavy chain V-III region BRO HV305_HUMAN 13 kDa P01766 Homo sapiens 50 Dynactin subunit 2 DCTN2_HUMAN 44 kDa Q13561 Homo sapiens 51 Inter-alpha-trypsin inhibitor heavy ITIH1_HUMAN 101 kDa P19827 Homo sapiens chain H1 52 Complement C2 CO2_HUMAN 83 kDa P06681 Homo sapiens 53 Serum amyloid P-component SAMP_HUMAN 25 kDa P02743 Homo sapiens 54 Lamin-A/C LMNA_HUMAN 74 kDa P02545 Homo sapiens 55 Phosphatidylinositol-glycan-specific PHLD1_HUMAN 92 kDa P80108 Homo sapiens phospholipase D 56 Alpha-1-acid glycoprotein 1 A1AG1_HUMAN 24 kDa P02763 Homo sapiens 57 Pigment epithelium-derived factor PEDF_HUMAN 46 kDa P36955 Homo sapiens 58 Apolipoprotein C-I APOC1_HUMAN 9 kDa P02654 Homo sapiens 59 Transcription intermediary factor 1- TIF1B_HUMAN 89 kDa Q13263 Homo sapiens beta 60 Leucine-rich alpha-2-glycoprotein A2GL_HUMAN 38 kDa P02750 Homo sapiens 61 Lumican LUM_HUMAN 38 kDa P51884 Homo sapiens 62 Ig kappa chain V-III region SIE KV302_HUMAN 12 kDa P01620 Homo sapiens 63 Actin, cytoplasmic 1 ACTB_HUMAN 42 kDa P60709 Homo sapiens 64 Apolipoprotein E APOE_HUMAN 36 kDa P02649 Homo sapiens 65 Kallistatin KAIN_HUMAN 49 kDa P29622 Homo sapiens 66 Myosin-15 MYH15_HUMAN 225 kDa Q9Y2K3 Homo sapiens 67 Carboxypeptidase B2 CBPB2_HUMAN 48 kDa Q96IY4 Homo sapiens 68 Serum amyloid A-4 protein SAA4_HUMAN 15 kDa P35542 Homo sapiens 69 Cholinesterase CHLE_HUMAN 68 kDa P06276 Homo sapiens 70 Apolipoprotein D APOD_HUMAN 21 kDa P05090 Homo sapiens 71 Tubulin beta-2C chain TBB2C_HUMAN 50 kDa P68371 Homo sapiens 72 Haptoglobin-related protein HPTR_HUMAN 39 kDa P00739 Homo sapiens 73 Hemoglobin subunit delta HBD_HUMAN 16 kDa P02042 Homo sapiens 74 Ig kappa chain V-IV region Len KV402_HUMAN 13 kDa P01625 Homo sapiens 75 Ig alpha-2 chain C region IGHA2_HUMAN 37 kDa P01877 Homo sapiens 76 Proteoglycan 4 PRG4_HUMAN 151 kDa Q92954 Homo sapiens 77 Histone H2A type 1-D H2A1D_HUMAN 14 kDa P20671 Homo sapiens 78 Biotinidase BTD_HUMAN 61 kDa P43251 Homo sapiens 79 Carboxypeptidase N catalytic chain CBPN_HUMAN 52 kDa P15169 Homo sapiens 80 Plasma kallikrein KLKB1_HUMAN 71 kDa P03952 Homo sapiens 81 Ig kappa chain V-II region TEW KV204_HUMAN 12 kDa P01617 Homo sapiens 82 Apolipoprotein C-II APOC2_HUMAN 11 kDa P02655 Homo sapiens 83 DNA-dependent protein kinase PRKDC_HUMAN 469 kDa P78527 Homo sapiens catalytic subunit 84 Heterogeneous nuclear HNRL1_HUMAN 96 kDa Q9BUJ2 Homo sapiens ribonucleoprotein U-like protein 1 85 Apolipoprotein B-100 APOB_HUMAN 516 kDa P04114 Homo sapiens 86 Heterogeneous nuclear ROA2_HUMAN 37 kDa P22626 Homo sapiens ribonucleoproteins A2/B1 87 Tubulin alpha-ubiquitous chain TBAK_HUMAN 50 kDa P68363 Homo sapiens 88 Gelsolin GELS_HUMAN 86 kDa P06396 Homo sapiens 89 Tetranectin TETN_HUMAN 23 kDa P05452 Homo sapiens 90 Alpha-1-acid glycoprotein 2 A1AG2_HUMAN 24 kDa P19652 Homo sapiens 91 Ig heavy chain V-III region GAL HV320_HUMAN 13 kDa P01781 Homo sapiens 92 IgGFc-binding protein FCGBP_HUMAN 572 kDa Q9Y6R7 Homo sapiens 93 Lactotransferrin TRFL_HUMAN 78 kDa P02788 Homo sapiens 94 Histone H4 H4_HUMAN 11 kDa P62805 Homo sapiens 95 Glutathione peroxidase 3 GPX3_HUMAN 26 kDa P22352 Homo sapiens 96 Histone H2B type 1-M H2B1M_HUMAN 14 kDa Q99879 Homo sapiens 97 Apolipoprotein M APOM_HUMAN 21 kDa O95445 Homo sapiens 98 E3 SUMO-protein ligase RanBP2 RBP2_HUMAN 358 kDa P49792 Homo sapiens 99 Ig kappa chain V-I region EU KV106_HUMAN 12 kDa P01598 Homo sapiens 100 Ig kappa chain V-I region Lay KV113_HUMAN 12 kDa P01605 Homo sapiens 101 Dermcidin OS = Homo sapiens DCD_HUMAN 11 kDa P81605 Homo sapiens 102 Leucine-rich repeats and LRIG2_HUMAN 119 kDa O94898 Homo sapiens immunoglobulin-like domains protein 2 103 Protein S100-A8 S10A8_HUMAN 11 kDa P05109 Homo sapiens 104 Glyceraldehyde-3-phosphate G3P_HUMAN 36 kDa P04406 Homo sapiens dehydrogenase 105 Myosin-9 MYH9_HUMAN 227 kDa P35579 Homo sapiens 106 Ig kappa chain V-III region VG KV309_HUMAN 13 kDa P04433 Homo sapiens (Fragment) 107 Monocyte differentiation antigen CD14_HUMAN 40 kDa P08571 Homo sapiens CD14 108 Heterogeneous nuclear HNRH1_HUMAN 49 kDa P31943 Homo sapiens ribonucleoprotein H 109 ATP-dependent DNA helicase 2 KU70_HUMAN 70 kDa P12956 Homo sapiens subunit 1 110 Protein TFG TFG_HUMAN 43 kDa Q92734 Homo sapiens 111 Coagulation factor XII FA12_HUMAN 68 kDa P00748 Homo sapiens 112 Filaggrin-2 FILA2_HUMAN 248 kDa Q5D862 Homo sapiens 113 Prothrombin THRB_HUMAN 70 kDa P00734 Homo sapiens 114 Sex hormone-binding globulin SHBG_HUMAN 44 kDa P04278 Homo sapiens 115 Heat shock cognate 71 kDa protein HSP7C_HUMAN 71 kDa P11142 Homo sapiens 116 Histidine-rich glycoprotein HRG_HUMAN 60 kDa P04196 Homo sapiens 117 Heterogeneous nuclear ROA3_HUMAN 40 kDa P51991 Homo sapiens ribonucleoprotein A3 118 Ig kappa chain V-III region B6 KV301_HUMAN 12 kDa P01619 Homo sapiens 119 Testis-expressed sequence 15 TEX15_HUMAN 315 kDa Q9BXT5 Homo sapiens protein 120 Polyadenylate-binding protein 1 PABP1_HUMAN 71 kDa P11940 Homo sapiens 121 Lipopolysaccharide-binding protein LBP_HUMAN 53 kDa P18428 Homo sapiens 122 Lamina-associated polypeptide 2, LAP2A_HUMAN 75 kDa P42166 Homo sapiens isoform alpha 123 Desmoplakin DESP_HUMAN 332 kDa P15924 Homo sapiens 124 Protein S100-A9 S10A9_HUMAN 13 kDa P06702 Homo sapiens 125 Far upstream element-binding FUBP2_HUMAN 73 kDa Q92945 Homo sapiens protein 2 126 Platelet glycoprotein Ib alpha chain GP1BA_HUMAN 69 kDa P07359 Homo sapiens 127 Zinc finger and SCAN domain- ZSC29_HUMAN 97 kDa Q8IWY8 Homo sapiens containing protein 29 128 Heterogeneous nuclear HNRH3_HUMAN 37 kDa P31942 Homo sapiens ribonucleoprotein H3 129 Histone H3.3 H33_HUMAN 15 kDa P84243 Homo sapiens 130 Stress-70 protein, mitochondrial GRP75_HUMAN 74 kDa P38646 Homo sapiens 131 Ig kappa chain V-I region AG KV101_HUMAN 12 kDa P01593 Homo sapiens 132 Elongation factor 1-alpha 1 EF1A1_HUMAN 50 kDa P68104 Homo sapiens 133 RNA-binding protein FUS FUS_HUMAN 53 kDa P35637 Homo sapiens 134 DNA repair and recombination RA54B_HUMAN 103 kDa Q9Y620 Homo sapiens protein RAD54B 135 Hornerin HORN_HUMAN 282 kDa Q86YZ3 Homo sapiens 136 Tubulin beta chain TBB5_HUMAN 50 kDa P07437 Homo sapiens 137 Heterogeneous nuclear HNRPC_HUMAN 34 kDa P07910 Homo sapiens ribonucleoproteins C1/C2 138 RecQ-mediated genome instability RMI1_HUMAN 70 kDa Q9H9A7 Homo sapiens protein 1 139 Desmocollin-1 DSC1_HUMAN 100 kDa Q08554 Homo sapiens 140 Xaa-Pro dipeptidase PEPD_HUMAN 55 kDa P12955 Homo sapiens 141 Junction plakoglobin PLAK_HUMAN 82 kDa P14923 Homo sapiens 142 U3 small nucleolar RNA-associated UT14C_HUMAN 87 kDa Q5TAP6 Homo sapiens protein 14 homolog C 143 Obscurin OBSCN_HUMAN 868 kDa Q5VST9 Homo sapiens 144 Elongation factor Tu, mitochondrial EFTU_HUMAN 50 kDa P49411 Homo sapiens 145 Heterogeneous nuclear HNRPF_HUMAN 46 kDa P52597 Homo sapiens ribonucleoprotein F 146 Desmoglein-1 DSG1_HUMAN 114 kDa Q02413 Homo sapiens 147 Heterogeneous nuclear ROA1_HUMAN 39 kDa P09651 Homo sapiens ribonucleoprotein Al 148 ELAV-like protein 1 ELAV1_HUMAN 36 kDa Q15717 Homo sapiens 149 Pyrroline-5-carboxylate reductase 1, P5CR1_HUMAN 33 kDa P32322 Homo sapiens mitochondrial 150 Ig lambda chain V-I region WAH LV106_HUMAN 12 kDa P04208 Homo sapiens 151 Zinc finger and BTB domain- ZBT10_HUMAN 92 kDa Q96DT7 Homo sapiens containing protein 10 152 Cystatin-C CYTC_HUMAN 16 kDa P01034 Homo sapiens 153 Major vault protein MVP_HUMAN 99 kDa Q14764 Homo sapiens 154 Phosphatidylcholine-sterol LCAT_HUMAN 50 kDa P04180 Homo sapiens acyltransferase 155 Sulfhydryl oxidase 1 QSCN6_HUMAN 83 kDa O00391 Homo sapiens 156 Far upstream element-binding FUBP3_HUMAN 62 kDa Q96I24 Homo sapiens protein 3 157 Ig delta chain C region IGHD_HUMAN 42 kDa P01880 Homo sapiens 158 Plasma serine protease inhibitor IPSP_HUMAN 46 kDa P05154 Homo sapiens 159 Heterogeneous nuclear HNRPR_HUMAN 71 kDa O43390 Homo sapiens ribonucleoprotein R 160 Hemoglobin subunit gamma-1 HBG1_HUMAN 16 kDa P69891 Homo sapiens (+1) 161 TAR DNA-binding protein 43 TADBP_HUMAN 45 kDa Q13148 Homo sapiens 162 Neural cell adhesion molecule L1- CHL1_HUMAN 135 kDa O00533 Homo sapiens like protein 163 Bifunctional polynucleotide PNKP_HUMAN 57 kDa Q96T60 Homo sapiens phosphatase/kinase 164 Fibrocystin PKHD1_HUMAN 447 kDa Q8TCZ9 Homo sapiens 165 Heterogeneous nuclear ROAA_HUMAN 36 kDa Q99729 Homo sapiens ribonucleoprotein A/B 166 Peroxiredoxin-1 PRDX1_HUMAN 22 kDa Q06830 Homo sapiens 167 40S ribosomal protein S16 RS16_HUMAN 16 kDa P62249 Homo sapiens 168 Heat shock 70 kDa protein 1 HSP71_HUMAN 70 kDa P08107 Homo sapiens 169 40S ribosomal protein S25 RS25_HUMAN 14 kDa P62851 Homo sapiens 170 Fetuin-B FETUB_HUMAN 42 kDa Q9UGM5 Homo sapiens 171 Transferrin receptor protein 1 TFR1_HUMAN 85 kDa P02786 Homo sapiens 172 Myelin basic protein MBP_HUMAN 33 kDa P02686 Homo sapiens 173 Aldo-keto reductase family 1 AK1C1_HUMAN 37 kDa Q04828 Homo sapiens member C1 174 Uncharacterized protein C19orf21 CS021_HUMAN 75 kDa Q8IVT2 Homo sapiens

175 TATA-binding protein-associated RBP56_HUMAN 62 kDa Q92804 Homo sapiens factor 2N 176 ATP-dependent DNA helicase 2 KU86_HUMAN 83 kDa P13010 Homo sapiens subunit 2 177 Sulfide:quinone oxidoreductase, SQRD_HUMAN 50 kDa Q9Y6N5 Homo sapiens mitochondrial 178 Interleukin enhancer-binding factor ILF2_HUMAN 43 kDa Q12905 Homo sapiens 2 179 Prostaglandin F2-alpha receptor PF2R_HUMAN 40 kDa P43088 Homo sapiens 180 Peptidase inhibitor 16 PI16_HUMAN 49 kDa Q6UXB8 Homo sapiens 181 Actin, alpha cardiac muscle 1 ACTC_HUMAN 42 kDa P68032 Homo sapiens 182 E3 SUMO-protein ligase RanBP2 RBP2_HUMAN 358 kDa P49792 Homo sapiens Mitochondrial import inner membrane translocase subunit 183 TIM50 TIM50_HUMAN 40 kDa Q3ZCQ8 Homo sapiens 184 Heterogeneous nuclear HNRPD_HUMAN 38 kDa Q14103 Homo sapiens ribonucleoprotein D0 185 Interleukin enhancer-binding factor ILF3_HUMAN 95 kDa Q12906 Homo sapiens 3 186 Heterogeneous nuclear HNRPG_HUMAN 42 kDa P38159 Homo sapiens ribonucleoprotein G 187 40S ribosomal protein S4, X isoform RS4X_HUMAN 30 kDa P62701 Homo sapiens 188 Caspase-14 CASPE_HUMAN 28 kDa P31944 Homo sapiens 189 Annexin A2 ANXA2_HUMAN 39 kDa P07355 Homo sapiens 190 Isocitrate dehydrogenase [NAD] IDH3A_HUMAN 40 kDa P50213 Homo sapiens subunit alpha, mitochondrial 191 Cysteine-rich secretory protein 3 CRIS3_HUMAN 28 kDa P54108 Homo sapiens 192 Myeloperoxidase PERM_HUMAN 84 kDa P05164 Homo sapiens 193 Transformation/transcription TRRAP_HUMAN 438 kDa Q9Y4A5 Homo sapiens domain-associated protein 194 Isocitrate dehydrogenase [NAD] IDH3B_HUMAN 42 kDa 043837 Homo sapiens subunit beta, mitochondrial 195 Collagen alpha-2(V) chain CO5A2_HUMAN 145 kDa P05997 Homo sapiens 196 Baculoviral IAP repeat-containing BIRC6_HUMAN 528 kDa Q9NR09 Homo sapiens protein 6 197 Bleomycin hydrolase BLMH_HUMAN 53 kDa Q13867 Homo sapiens 198 Poly(rC)-binding protein 1 PCBP1_HUMAN 37 kDa Q15365 Homo sapiens 199 Tubulin beta-3 chain TBB3_HUMAN 50 kDa Q13509 Homo sapiens 200 60 kDa heat shock protein, CH60_HUMAN 61 kDa P10809 Homo sapiens mitochondrial 201 ATP synthase subunit beta, ATPB_HUMAN 57 kDa P06576 Homo sapiens mitochondrial 202 40S ribosomal protein S18 RS18_HUMAN 18 kDa P62269 Homo sapiens 203 DnaJ homolog subfamily A member DNJA1_HUMAN 45 kDa P31689 Homo sapiens 1 204 RNA-binding motif, single-stranded- RBMS1_HUMAN 45 kDa P29558 Homo sapiens interacting protein 1 205 40S ribosomal protein S19 RS19_HUMAN 16 kDa P39019 Homo sapiens 206 Tubulin alpha-1 chain TBA1_HUMAN 50 kDa P68366 Homo sapiens 207 Protein Hook homolog 1 HOOK1_HUMAN 85 kDa Q9UJC3 Homo sapiens 208 Brother of CDO BOC_HUMAN 121 kDa Q9BWV1 Homo sapiens 209 Signal recognition particle 54 kDa SRP54_HUMAN 56 kDa P61011 Homo sapiens protein 210 Trinucleotide repeat-containing 6B TNC6B_HUMAN 194 kDa Q9UPQ9 Homo sapiens protein 211 Aldehyde dehydrogenase, dimeric AL3A1_HUMAN 50 kDa P30838 Homo sapiens NADP-preferring 212 40S ribosomal protein S3 RS3_HUMAN 27 kDa P23396 Homo sapiens 213 Fatty aldehyde dehydrogenase AL3A2_HUMAN 55 kDa P51648 Homo sapiens 214 Electron transfer flavoprotein ETFA_HUMAN 35 kDa P13804 Homo sapiens subunit alpha, mitochondrial 215 RNA-binding protein EWS EWS_HUMAN 68 kDa Q01844 Homo sapiens 216 Histone H2A.V H2AV_HUMAN 14 kDa Q71UI9 Homo sapiens Heterogeneous nuclear (+1) 217 ribonucleoprotein Q HNRPQ_HUMAN 70 kDa O60506 Homo sapiens 218 Pyrroline-5-carboxylate reductase P5CR2_HUMAN 34 kDa Q96C36 Homo sapiens 219 Prohibitin PHB_HUMAN 30 kDa P35232 Homo sapiens 220 RuvB-like 1 RUVB1_HUMAN 50 kDa Q9Y265 Homo sapiens 221 Protein-glutamine gamma- TGM2_HUMAN 77 kDa P21980 Homo sapiens glutamyltransferase 222 Ubiquitin-associated protein 2 UBAP2_HUMAN 117 kDa Q5T6F2 Homo sapiens 223 Latent-transforming growth factor LTB1L_HUMAN 187 kDa Q14766 Homo sapiens beta-binding protein 1 224 Thioredoxin domain-containing TXND2_HUMAN 60 kDa Q86VQ3 Homo sapiens protein 2 225 Leucine-rich repeat-containing LRC17_HUMAN 52 kDa Q8N6Y2 Homo sapiens protein 17 226 Protein FAM105B F105B_HUMAN 40 kDa Q96BN8 Homo sapiens 227 Myosin-Ia MYO1A_HUMAN 118 kDa Q9UBC5 Homo sapiens Ras GTPase-activating-like protein 228 IQGAP2 IQGA2_HUMAN 181 kDa Q13576 Homo sapiens 229 26Sproteasome non-ATPase PSMD1_HUMAN 102 kDa Q99460 Homo sapiens regulatory subunit 1 230 Actin, alpha skeletal muscle ACTS_HUMAN 42 kDa P68133 Homo sapiens 231 Cleavage stimulation factor subunit CSTF3_HUMAN 83 kDa Q12996 Homo sapiens 3 232 Clusterin CLUS_HUMAN 52 kDa P10909 Homo sapiens 233 Fibrocystin-L PKHL1_HUMAN 466 kDa Q86WI1 Homo sapiens 234 InaD-like protein INADL_HUMAN 196 kDa Q8NI35 Homo sapiens 235 Keratin, type I cuticular Ha1 K1H1_HUMAN 47 kDa Q15323 Homo sapiens 236 Keratin, type I cuticular Ha2 K1H2_HUMAN 50 kDa Q14532 Homo sapiens 237 Keratin, type I cuticular Ha6 K1H6_HUMAN 52 kDa O76013 Homo sapiens 238 Keratin, type I cytoskeletal 10 K1C10_HUMAN 59 kDa P13645 Homo sapiens 239 Keratin, type I cytoskeletal 14 K1C14_HUMAN 52 kDa P02533 Homo sapiens 240 Keratin, type I cytoskeletal 16 K1C16_HUMAN 51 kDa P08779 Homo sapiens 241 Keratin, type I cytoskeletal 18 K1C18_HUMAN 48 kDa P05783 Homo sapiens 242 Keratin, type I cytoskeletal 19 K1C19_HUMAN 44 kDa P08727 Homo sapiens 243 Keratin, type I cytoskeletal 9 K1C9_HUMAN 62 kDa P35527 Homo sapiens 244 Keratin, type II cuticular Hb1 KRT81_HUMAN 55 kDa Q14533 Homo sapiens 245 Keratin, type II cuticular Hb5 KRT85_HUMAN 56 kDa P78386 Homo sapiens 246 Keratin, type II cytoskeletal 1 K2C1_HUMAN 66 kDa P04264 Homo sapiens 247 Keratin, type II cytoskeletal 1b K2C1B_HUMAN 62 kDa Q7Z794 Homo sapiens 248 Keratin, type II cytoskeletal 2 K22E_HUMAN 65 kDa P35908 Homo sapiens epidermal 249 Keratin, type II cytoskeletal 4 K2C4_HUMAN 57 kDa P19013 Homo sapiens 250 Keratin, type II cytoskeletal 5 K2C5_HUMAN 62 kDa P13647 Homo sapiens 251 Keratin, type II cytoskeletal 6B K2C6B_HUMAN 60 kDa P04259 Homo sapiens 252 Keratin, type II cytoskeletal 7 K2C7_HUMAN 51 kDa P08729 Homo sapiens 253 Keratin, type II cytoskeletal 8 K2C8_HUMAN 54 kDa P05787 Homo sapiens 254 Lymphoid-restricted membrane LRMP_HUMAN 62 kDa Q12912 Homo sapiens protein 255 Myosin light chain 1, skeletal muscle MLE1_HUMAN 21 kDa P05976 Homo sapiens isoform 256 Myosin-2 MYH2_HUMAN 223 kDa Q9UKX2 Homo sapiens 257 Protein Shroom3 SHRM3_HUMAN 216 kDa Q8TF72 Homo sapiens 258 Ras-related GTP-binding protein A RRAGA_HUMAN 37 kDa Q7L523 Homo sapiens 259 Sushi, nidogen and EGF-like SNED1_HUMAN 152 kDa Q8TER0 Homo sapiens domain-containing protein 1 260 Retinol-binding protein 4 RET4_HUMAN 23 kDa P02753 Homo sapiens

TABLE-US-00002 TABLE 2 Proteins identified in spent media of human embryos. * SEQID Protein Molecular Accession Day of Embryo Nature of NO. Protein Name Entry Name Weight Number Development Viability Marker 261 14-3-3 protein zeta/delta 1433Z_HUMAN 28 kDa P63104 D 1 262 6-phosphofructokinase, muscle K6PF_HUMAN 85 kDa P08237 D 1 type, isoform 1 263 6-phosphofructokinase, muscle K6PF_HUMAN 82 kDA P08237-2 D 1 type, isoform 1 264 Adenylate kinase isoenzyme 1 KAD1_HUMAN 22 kDa P00568 D 1 265 Alpha-actinin-2 ACTN2_HUMAN 104 kDa P35609 D 1 266 Beta-enolase, isoform 1 ENOB_HUMAN 47 kDa P13929 D 1 267 Beta-enolase, isoform 2 ENOB_HUMAN 44 kDa P13929-2 D 1 268 Beta-enolase, isoform 3 ENOB_HUMAN 42 kDa P13929-3 D 1 269 Creatine kinase M-type KCRM_HUMAN 43 kDa P06732 D 1 270 Fructose-bisphosphate aldolase A ALDOA_HUMAN 39 kDa P04075 D 1 271 Glycogen phosphorylase, muscle form PYGM_HUMAN 97 kDa P11217 D 1 272 L-lactate dehydrogenase A chain LDHA_HUMAN 37 kDa P00338 D 1 273 Myosin-1 MYH1_HUMAN 223 kDa P12882 D 1 274 Myosin-2 MYH2_HUMAN 223 kDa Q9UKX2 D 1 275 Myosin-3 MYH3_HUMAN 224 kDa P11055 D 1 276 Myosin-4 MYH4_HUMAN 223 kDa Q9Y623 D 1 277 Myosin-6 MYH6_HUMAN 224 kDa P13533 D 1 278 Myosin-7 MYH7_HUMAN 223 kDa P12883 D 1 279 Myosin-8 MYH8_HUMAN 223 kDa P13535 D 1 280 Phosphoglucomutase-1, isoform 1 PGM1_HUMAN 61 kDa P36871 D 1 281 Phosphoglucomutase-1, isoform 2 PGM1_HUMAN 64 kDa P36871-2 D 1 282 Phosphoglycerate kinase 1 PGK1_HUMAN 45 kDa P00558 D 1 283 Phosphoglycerate mutase 1 PGAM1_HUMAN 29 kDa P18669 D 1 284 Pyruvate kinase isozyme M2 KPYM_HUMAN 58 kDa P14618 D 1 285 Pyruvate kinase isozyme M1 KPYM_HUMAN 58 kDa P14618-2 D 1 286 Sarcoplasmic/endoplasmic reticulum AT2A1_HUMAN 110 kDa O14983 D 1 calcium ATPase 1, Isoform SERCA1B 287 Sarcoplasmic/endoplasmic reticulum AT2A1_HUMAN 109 kDa O14983-2 D 1 calcium ATPase 1, Isoform SERCA1A 288 Sarcoplasmic/endoplasmic reticulum AT2A2_HUMAN 115 kDa P16615 D 1 calcium ATPase 2, isoform 1 289 Sarcoplasmic/endoplasmic reticulum AT2A2_HUMAN 110 kDa P16615-2 D 1 calcium ATPase 2, isoform 2 290 Sarcoplasmic/endoplasmic reticulum AT2A2_HUMAN 110 kDa P16615-3 D 1 calcium ATPase 2, isoform 3 291 Sarcoplasmic/endoplasmic reticulum AT2A2_HUMAN 112 kDa P16615-4 D 1 calcium ATPase 2, isoform 4 292 Sarcoplasmic/endoplasmic reticulum AT2A2_HUMAN 110 kDa P16615-5 D 1 calcium ATPase 2, isoform 5 293 Triosephosphate isomerase, isoform 1 TPIS_HUMAN 27 kDa P60174 D 1 294 Triosephosphate isomerase, isoform 2 TPIS_HUMAN 27 kDa P60174-2| D 1 295 Tropomyosin alpha-1 chain, isoform 1 TPM1_HUMAN 33 kDa P09493 D 1 296 Tropomyosin alpha-1 chain, isoform 2 TPM1_HUMAN 27 kDa P09493-2 D 1 297 Tropomyosin alpha-1 chain, isoform 3 TPM1_HUMAN 33 kDa P09493-3 D 1 298 Tropomyosin alpha-1 chain, isoform 4 TPM1_HUMAN 33 kDa P09493-4 D 1 299 Tropomyosin alpha-1 chain, isoform 5 TPM1_HUMAN 28 kDa P09493-6 D 1 300 Tropomyosin alpha-3 chain, isoform 1 TPM3_HUMAN 33 kDa P06753 D 1 301 Tropomyosin alpha-3 chain, isoform 2 TPM3_HUMAN 29 kDa P06753-2 D 1 302 Tropomyosin alpha-3 chain, isoform 3 TPM3_HUMAN 29 kDa P06753-3 D 1 303 Collagen alpha-1(I) chain CO1A1_HUMAN 139 kDa P02452 D 1/D 3 304 Cystatin-A CYTA_HUMAN 11 kDa P01040 D 1/D 3 305 Filaggrin FILA_HUMAN 435 kDa P20930 D 1/D 3 306 Keratinocyte proline-rich protein KPRP_HUMAN 64 kDa Q5T749 D 1/D 3 307 Selenium-binding protein 1, isoform 1 SBP1_HUMAN 52 kDa Q13228 D 3 308 Selenium-binding protein 1, isoform 2 SBP1_HUMAN 45 kDa Q13228-2 D 3 309 Fatty acid-binding protein, epidermal FABP5_HUMAN 15 kDa Q01469 D 1/D 3/D 5 310 Galectin-7 LEG7_HUMAN 15 kDa P47929 D 1/D 3/D 5 Increased with implantation Success 311 Ubiquitin UBIQ_HUMAN X kDa P02248 D 1/D 3/D 5 Increased with implantation Success 312 Calmodulin-like protein 5 CALL5_HUMAN 16 kDa Q9NZT1 D 1/D 3/D 5 313 14-3-3 protein sigma 1433S_HUMAN 28 kDa P31947 D 1/D 3/D 5 Increased with implantation Success 314 Histone H2A type 1-A H2A1A_HUMAN 14 kDa Q96QV6 D 1/D 3/D 5 315 Histone H3.1t H31T_HUMAN 16 kDa Q16695 D 1/D 3/D 5 316 Histone H1.3 H13_HUMAN 22 kDa P16402 D 3/D 5 317 Lysozyme C LYSC_HUMAN 17 kDa P61626 D 3/D 5 Increased with Aneuploidy 318 Plakophilin-1, isoform 2 PKP1_HUMAN 83 kDa Q13835 D 3/D 5 Increased with implantation Success 319 Plakophilin-1, isoform 1 PKP1_HUMAN 80 kDa Q13835-2 D 3/D 5 Increased with implantation Success 320 Abhydrolase domain-containing AB12B_HUMAN 41 kDa Q7Z5M8 D 3/D 5 protein 12B, isoform 1 321 Abhydrolase domain-containing AB12B_HUMAN 32 kDa Q7Z5M8-2 D 3/D 5 protein 12B, isoform 2 322 Abhydrolase domain-containing AB12B_HUMAN 29 kDa Q7Z5M8-3 D 3/D 5 protein 12B, isoform 3 323 Abhydrolase domain-containing AB12B_HUMAN 13 kDa Q7Z5M8-4 D 3/D 5 protein 12B, isoform 4 324 Abhydrolase domain-containing AB12B_HUMAN 25 kDa Q7Z5M8-5 D 3/D 5 protein 12B, isoform 5 325 40S ribosomal protein SA RSSA_HUMAN 33 kDa P08865 D 5 326 Arginase-1, isoform 1 ARGI1_HUMAN 35 kDa P05089 D 5 327 Arginase-1, isoform 2 ARGI1_HUMAN 36 kDa P05089-2 D 5 328 Arginase-1, isoform 3 ARGI1_HUMAN 25 kDa P05089-3 D 5 329 Beta-2-microglobulin B2MG_HUMAN 14 kDa P61769 D 5 330 Carbonic anhydrase 2 CAH2_HUMAN 29 kDa P00918 D 5 331 Carboxypeptidase A4 CBPA4_HUMAN 47 kDa Q9UI42 D 5 332 Catalase CATA_HUMAN 60 kDa P04040 D 5 333 Cathepsin D CATD_HUMAN 45 kDa P07339 D 5 334 CD44 antigen, isoform 1 CD44_HUMAN 81 kDa P16070 D 5 Increased with implantation Success 335 CD44 antigen, isoform 2 CD44_HUMAN 3 kDa P16070-2 D 5 Increased with implantation Success 336 CD44 antigen, isoform 3 CD44_HUMAN 78 kDa P16070-3 D 5 Increased with implantation Success 337 CD44 antigen, isoform 4 CD44_HUMAN 77 kDa P16070-4 D 5 Increased with implantation Success 338 CD44 antigen, isoform 5 CD44_HUMAN 81 kDa P16070-5 D 5 Increased with implantation Success 339 CD44 antigen, isoform 6 CD44_HUMAN 77 kDa P16070-6 D 5 Increased with implantation Success 340 CD44 antigen, isoform 7 CD44_HUMAN 78 kDa P16070-7 D 5 Increased with implantation Success 341 CD44 antigen, isoform 8 CD44_HUMAN 74 kDa P16070-8 D 5 Increased with implantation Success 342 CD44 antigen, isoform 9 CD44_HUMAN 74 kDa P16070-9 D 5 Increased with implantation Success 343 CD44 antigen, isoform 10 CD44_HUMAN 53 kDa P16070-10 D 5 Increased with implantation Success 344 CD44 antigen, isoform 11 CD44_HUMAN 47 kDa P16070-11 D 5 Increased with implantation Success 345 CD44 antigen, isoform 12 CD44_HUMAN 39 kDa P16070-12 D 5 Increased with implantation Success 346 CD44 antigen, isoform 13 CD44_HUMAN 46 kDa P16070-13 D 5 Increased with implantation Success 347 CD44 antigen, isoform 14 CD44_HUMAN 43 kDa P16070-14 D 5 Increased with implantation Success 348 CD44 antigen, isoform 15 CD44_HUMAN 32 kDa P16070-15 D 5 Increased with implantation Success 349 CD44 antigen, isoform 16 CD44_HUMAN 73 kDa P16070-16 D 5 Increased with implantation Success 350 CD44 antigen, isoform 17 CD44_HUMAN 76 kDa P16070-17 D 5 Increased with implantation Success 351 Creatine kinase B-type KCRB_HUMAN 43 kDa P12277 D 5 352 Dermokine, isoform 1 DMKN_HUMAN 47 kDa Q6E0U4 D 5 353 Dermokine, isoform 2 DMKN_HUMAN 47 kDa Q6E0U4-2 D 5 354 Dermokine, isoform 3 DMKN_HUMAN 37 kDa Q6E0U4-3 D 5 355 Dermokine, isoform 4 DMKN_HUMAN 45 kDa Q6E0U4-4 D 5 356 Dermokine, isoform 5 DMKN_HUMAN 38 kDa Q6E0U4-5 D 5 357 Dermokine, isoform 6 DMKN_HUMAN 42 kDa Q6E0U4-6 D 5 358 Dermokine, isoform 7 DMKN_HUMAN 35 kDa Q6E0U4-7 D 5 359 Dermokine, isoform 8 DMKN_HUMAN 20 kDa Q6E0U4-8 D 5 360 Dermokine, isoform 9 DMKN_HUMAN 19 kDa Q6E0U4-9 D 5 361 Dermokine, isoform 10 DMKN_HUMAN 17 kDa Q6E0U4-10 D 5 362 Dermokine, isoform 11 DMKN_HUMAN 15 kDa Q6E0U4-11 D 5 363 Dermokine, isoform 12 DMKN_HUMAN 16 kDa Q6E0U4-12 D 5 364 Dermokine, isoform 13 DMKN_HUMAN 14 kDa Q6E0U4-13 D 5 365 Dermokine, isoform 14 DMKN_HUMAN 15 kDa Q6E0U4-14 D 5 366 Dermokine, isoform 15 DMKN_HUMAN 10 kDa Q6E0U4-15 D 5 367 Extracellular glycoprotein lacritin LACRT_HUMAN 14 kDa Q9GZZ8 D 5 368 Ferritin light chain (Ferritin L FRIL_HUMAN 20 kDa P02792 D 5 Increased with subunit) implantation Success 369 Fibrinogen gamma chain, isoform FIBG_HUMAN 52 kDa P02679 D 5 gamma-B 370 Fibrinogen gamma chain, isoform FIBG_HUMAN 49 kDa P02679-2 D 5 gamma-A 371 Gamma-enolase ENOG_HUMAN 47 kDa P09104 D 5 Increased with implantation Success 372 Gamma-glutamylcyclotransferase, GGCT_HUMAN 21 kDa O75223 D 5 isoform 1 373 Gamma-glutamylcyclotransferase, GGCT_HUMAN 13 kDa O75223-2 D 5 isoform 2 374 Heat shock protein beta-1 HSPB1_HUMAN 23 kDa P04792 D 5 Increased with implantation Success 375 Histidine ammonia-lyase HUTH_HUMAN 73 kDa P42357 D 5 376 Ig mu chain C region, isoform 1 IGHM_HUMAN 49 kDa P01871 D 5 377 Ig mu chain C region, isoform 2 IGHM_HUMAN 52 kDa P01871-2 D 5 378 Interleukin-1 family member 7 IL1F7_HUMAN 224 kDa Q9NZH6 D 5 precursor, isoform B 379 Interleukin-1 family member 7 IL1F7_HUMAN 22 kDa Q9NZH6-2 D 5 precursor, isoform A 380 Interleukin-1 family member 7 IL1F7_HUMAN 20 kDa Q9NZH6-2 D 5 precursor, isoform C 381 Interleukin-1 family member 7 IL1F7_HUMAN 22 kDa Q9NZH6-2 D 5 precursor, isoform D 382 Interleukin-1 family member 7 IL1F7_HUMAN 17 kDa Q9NZH6-2 D 5 precursor, isoform E 383 Lipocalin-1 LCN1_HUMAN 19 kDa P31025 D 5 Increased with Aneuploidy 384 Methionine synthase METH_HUMAN 141 kDa Q99707 D 5 385 Peroxiredoxin-2 PRDX2_HUMAN 22 kDa P32119 D 5 386 Proprotein convertase PCSK9_HUMAN 74 kDa Q8NBP7 D 5 subtilisin/kexin type 9, isoform 1 387 Proprotein convertase PCSK9_HUMAN 21 kDa Q8NBP7-2 D 5 subtilisin/kexin type 9, isoform 2 388 Protein POF1B, isoform 1 POF1B_HUMAN 69 kDa Q8WVV4 D 5 389 Protein POF1B, isoform 2 POF1B_HUMAN 68 kDa Q8WVV4-2 D 5 390 Protein POF1B, isoform 3 POF1B_HUMAN 35 kDa Q8WVV4-3 D 5 391 Protein S100-A7 S10A7_HUMAN 11 kDa P31151 D 5 Increased with implantation Success 392 Protein-glutamine gamma- TGM3_HUMAN 77 kDa Q08188 D 5 glutamyltransferase E 393 Pyruvate kinase isozymes M1/M2, KPYM_HUMAN 58 kDa P14618 D 5 isoform M2 394 Pyruvate kinase isozymes M1/M2, KPYM_HUMAN 58 kDa P14618-2 D 5 isoform M1 395 Receptor-type tyrosine-protein PTPRG_HUMAN 162 kDa P23470 D 5 phosphatase gamma, isoform 1 396 Receptor-type tyrosine-protein PTPRG_HUMAN 159 kDa P23470-2 D 5 phosphatase gamma, isoform 2 397 Serine protease inhibitor Kazal-type ISK5_HUMAN 121 kDa Q9NQ38 D 5 Increased with 5, isoform f1 implantation Success 398 Serine protease inhibitor Kazal-type ISK5_HUMAN 104 kDa Q9NQ38 D 5 Increased with 5, isoform short implantation Success 399 Serine protease inhibitor Kazal-type ISK5_HUMAN 124 kDa Q9NQ38 D 5 Increased with

5, isoform long implantation Success 400 Serpin B3, isoform 1 SPB3_HUMAN 45 kDa P29508 D 5 401 Serpin B3, isoform 2 SPB3_HUMAN 39 kDa P29508-2 D 5 402 Uteroglobin UTER_HUMAN 10 kDa P11684 D 5 Increased with implantation Success 403 V-set and immunoglobulin domain- VSIG8_HUMAN 44 kDa Q5VU13 D 5 Increased with containing protein 8, isoform 1 implantation Success 404 V-set and immunoglobulin domain- VSIG8_HUMAN 25 kDa D 5 Increased with containing protein 8, isoform 1 implantation Success * All Proteins are from Homo Sapiens

Example 2

Secretome Profiling to Predict Implantation Potential of a Human Embryo

[0047] The following nonlimiting example teaches by way of illustration, not by limitation, the fabrication and employment of a customized immunoassay test kit for secretome profiling of a human embryo. Immunoassay test kit fabrication occurs by modifying the well surfaces of a 96-well microtiter plate. Each separate well of the microtiter plate is incubated with an unlabelled capture antibody that recognizes one specific protein from the secretome panel. Custom capture antibodies are purchased from Rockland Immunochemicals, Inc. After incubation for 12 hours at 4.degree. C., capture antibodies passively adsorb to the well surface. Subsequently, all wells of the 96-well plate are washed three times with a buffer comprising 1.times. phosphate buffer saline and 0.1% Tween-20 (PBST), blocked with 1% bovine serum albumin (BSA) for 1 hour at ambient temperature and washed three times with PBST.

[0048] Human cleavage-stage embryos are cultured in 10 .mu.L drops of G1 supplemented with 2.5 mg/mL recombinant albumin under oil at 37.degree. C., 6% CO.sub.2, 5% O.sub.2 for 24 hours. The embryos are washed twice in G2 culture media and further cultured in 10 .mu.L, drops of G2 supplemented with 2.5 mg/mL recombinant albumin under oil at 37.degree. C., 6% CO.sub.2, 5% O.sub.2 for 48 hours with a fresh drop of G2 media added after 24 hours. Spent media samples of blastocysts are transferred into 0.65 mL Eppendorf tubes. Control groups comprise media cultured and collected under the same conditions but without embryos.

[0049] Secretome profiling proceeds using spent media culture in a sandwich ELISA format. The spent media culture is diluted into phosphate buffer saline (PBS) buffer and further distributed to separate wells of said customized microtiter plate. After incubation for one hour at room temperature, the samples are aspirated and all wells are washed four times with PBST buffer. Primary antibodies, each recognizing a distinct protein from the secretome panel, are distributed into wells that correspond to appropriate capture antibodies and permitted to incubate for one hour at room temperature. The wells are washed four times with PBST and incubated with a secondary antibody conjugated with an enzyme. The term "secondary antibody" refers to an antibody that binds to primary antibodies and may be conjugated with detection probes such as enzymes or fluorophores. In one embodiment, the secondary antibody contains the horseradish peroxidase (HRP) enzyme that converts the chromogenic substrate, 3,3',5,5'-tetramethylbenzidene (TMB), into a blue product quantified at 655 nm on a spectrophotometer. In another embodiment, the HRP enzymatic reaction is stopped with a solution containing sulfuric acid and quantified at 450 nm.

[0050] In one embodiment, secretome profiling of a secretome panel of 261 individual proteins (Table 1), each selected for significant developmental competence and implantation potential, are monitored using said ELISA immunoassay test kit. In another embodiment, secretome profiling of a secretome panel comprising 37 proteins (Table 2) are monitored using said ELISA immunoassay test kit. Secretome profiling of the 37 individual proteins (Table 2) from spent media of human embryos correlates with embryonic viability and euploidy.

[0051] Changes may be made in the above methods and systems without departing from the scope hereof. It should be noted that the matter contained in the above description or shown in the accompanying drawings should be interpreted as illustrative and not in a limiting sense. The following claims are intended to cover all generic and specific features described herein, as well as all statements of the scope of the present method and system and reasonable variations thereof, which, as a matter of language, might be said to fall therebetween.

Sequence CWU 0 SQTB SEQUENCE LISTING The patent application contains a lengthy "Sequence Listing" section. A copy of the "Sequence Listing" is available in electronic form from the USPTO web site (http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20110245592A1). An electronic copy of the "Sequence Listing" will also be available from the USPTO upon request and payment of the fee set forth in 37 CFR 1.19(b)(3).

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