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Development of human embryo viability biomarker diagnostics for elective embryo transfer to improve pregnancy outcome: potential application in ART clinic

Primary Information



Project No.


Sanction and Project Initiation

Sanction No: F.No-35-10/2016-TS-I

Sanction Date: 27/02/2017

Project Initiation date: 08/02/2017

Project Duration: 36

Partner Ministry/Agency/Industry

Indian Council of Medical Research


Role of partner: 1. MHRD (Financial support) 2. ICMR (Financial support)


Support from partner: Financial support.

Principal Investigator

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Prof. Polani B. Seshagiri
Dept. of Molecular Reproduction, Development and Genetics, Indian Institute of Science, Bangalore

Host Institute


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Prof. Satish K. Adiga
Kasturba Medical College, Manipal University, Manipal

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Dr. S S Vasan
Manipal Ankur Andrology & Reproductive Services, Bangalore

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Dr. Varsha Samson Roy
Advanced Fertility Centre, Bangalore

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Dr. Geetanajali Sachdeva
National Institute for Research in Reproductive Health (ICMR), Parel, Mumbai 400012.


Scope and Objectives

To treat human infertility, significant advances have been made in the assisted reproductive technology (ART) sectors. Yet, the live-birth rate is < 15%. Among other causes, poor biological viability of embryos substantially contributes to pregnancy loss. Hence, there is an urgent unmet need to assess quality-viability of embryos, by a non invasive approach, by analysing embryo derived biomarkers in the spent medium. As depicted in the working hypothesis model (Fig. 1), this study aims to develop targeted immune-diagnostics based on HLA-G or and ubiquitin and potential biomarkers by non-targeted embryomics (Proteome and metabolome) technology platforms. It is envisaged that a point of care (PoC) molecular diagnostics could be potentially developed. This could be implemented, as an adjunct to embryo-score criteria, in ART clinics. Goals expected to be achieved are
(1) Development of immunodiagnostics for embryo biomarkers sHLA G & ubiquitin in spent medium: correlation with embryo quality score and pregnancy outcome.
(2) Embryomics analysis for embryo-viability biomarker discovery and its validation
(3) Retrospective correlative analysis between biomarkers prediction potential and pregnancy outcomes
(4) Development of POC molecular diagnostics platform kit for ART Clinic.


So far we successfully (a) measured sHLA G in ESM samples which showed embryo developmental stage correlation, albeit, with variability across clinics and (b) developed metabolomics profile with at least nine metabolites signature, associated with developing blastocysts which underwent ETs. After performing retrospective correlative analysis between biomarkers prediction potential with the embryo score and pregnancy outcome a point of care (PoC) molecular diagnostics could be potentially developed. This could be an adjunct to embryo score criteria, adopted in ART clinics. Human embryo viability discovery research could help in strategic improvement in pregnancy outcomes for infertile women.


Scientific Output

Human embryo spent medium samples were collected based on the experimental strategy depicted in Fig1. So far we have collected 3649 ESMs samples which include 1459 1162 and 1028 from KMC MAARS and AFC clinics, respectively. Pertaining to objectives 1 and 2 we have the following result: standardized the ELISA system to measure sHLA G and ubiquitin in ESM samples and observed development associated changes in sHLAG content with variability within and across clinics also the sHLAG concentration was different between ESMs of cleavage stage vs blastocysts samples for the freshET category (Fig 2) and for the frozenET category (Fig 3) Also, we observed a distinct nine metabolites signature in blastocyst ESM samples visavis control of the nine, seven metabolites were higher in ESMs visa vs control. Possible association of embryo metabolites profile with embryo quality scores and pregnancy outcome is to be established.


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Results and outcome till date

Progress made with respect to objective 1 Human embryo spent medium samples (hE SMs) were collected based on the experimental strategy depicted in Fig 1 Since the last month report update, we currently have procured additional 242 ES Ms (total so far 3953) from three participating ART clinics. In the last month, we have analysed about 170 E SMs (a total of 710) for embryo viability biomarker sHLA G and all estimations were Completed. of the 710 ESMs, we have analysed a total of 539 (223 fresh ETs and 316 frozen ETs) ESMs for embryo viability marker sHLA G. However, the association between pregnancy outcome and sHLAG levels is in progress. The data on the farmer aspect are procured from the participating ART clinics. As indicated in the last month report, in the current month, we have initiated the ELISA based estimations for human ubiquitin for E SMs of fresh ET and frozen ET samples. The pending E SM samples for ubiquitin estimation are to be performed once the ordered ELISA Kits is received. Progress made with respect to objective 2 As indicated in the last month report, we now have completed the metabolomics analysis of the eleven ESMs of non transferred blastocyst stage embryos and compared them with their respective control medium samples. For metabolomics analysis, we used both waters and luna columns (positive and negative modes). The analysis was carried out at an FDR of 0.25. When we compared the metabolic footprint or signature of the ESMs of the blastocyst stage embryos, We observed a distinct 4 metabolites signature in blastocyst ESM samples vis a vs control (Fig. 1) of the 4, three metabolites were significantly higher in ESMs vis a vs control medium. The identified metabolites were sarcosine, Glutamine, hydroxyisocaproic acid, shikimate (Fig 1). In the future we wish to perform a metabolomics analysis of more number of ESMs of blastocyst (Fresh and frozen ET category). Further, we will establish a possible association of embryo metabolite profile with embryo quality scores/pregnancy outcome, once the clinical data from the respective (participating) ART Clinics are received (is in progress).


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Societal benefit and impact anticipated

We have embarked on embryo biomarker discovery and molecular diagnostic kit development program for potential POC use in ART clinics.

Next steps

We will establish a possible association between embryo biomarkers and embryo quality scores and pregnancy outcome, once the clinical data from the respective (participating) ART clinics are received (in progress).

Publications and reports

A manuscript entitled, sHLA G levels and their significance in fresh and frozen ETs with respect to pregnancy outcome is being prepared for submission to a core journal related to reproductive medicine.



Scholars and Project Staff

Research Associate
1. Dr. Vani V (July 2017-ongoing)

Project Assistant
1. Ms. Akshatha D.P (Sept. 2017-ongoing)
2. Ms. Sheeba Lobo, Project trainee (Dec. 2017-ongoing)
3. Ms. Sai Prashanth S (May. 2019-ongoing)

Ph.D student
1. Ms. Sharmeli Sarkar (July 2016-ongoing)

Other information

Actual expenditure is shown here is for the duration of April to sept. 2019 and is based on new SAP Accounting system.

Financial Information

  • Total sanction: Rs. 19089600

  • Amount received: Rs. 10745100

  • Amount utilised for Equipment: Rs. 2000000

  • Amount utilised for Manpower: Rs. 538000

  • Amount utilised for Consumables: Rs. 386018

  • Amount utilised for Contingency: Rs. 184721

  • Amount utilised for Travel: Rs. 60515

  • Amount utilised for Other Expenses: 0

  • Amount utilised for Overheads: Rs. 2918400

Equipment and facilities


2. -80 degree Deep freezer (Eppendorf)

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