What the EPF?

We’ve Known How to Detect Fertilization Within 48 Hours Using Early Pregnancy Factor Since 1974. We Just Don’t

Determining when a cow is pregnant is important.

Can we all agree?

Determining when they die is also important.

According to one author, “Early embryonic death of calves due to sub-fertility in cows is of great economic concern to dairy industry.”¹

These three articles were published between 2021 and 2024 and focus on developing new ways to detect embryonic death earlier in cows.

• “Recent Possibilities for the Diagnosis of Early Pregnancy and Embryonic Mortality in Dairy Cows”
• “Non-invasive methods for diagnosing pregnancy in cows and their real value”
• “Selection of early pregnancy specific proteins and development a rapid immunochromatographic test strip in cows”

The researchers are investigating multiple approaches including measuring levels of interferon stimulated genes (ISGs), Toll-like receptor (TLRs), complement components, and pregnancy-associated plasma protein A (PAPPA) proteins.

They’re also measuring Early Pregnancy Factor (EPF).

Early Pregnancy Factor

EPF is a heat-shock protein (HSP) referred to as HSP-10 or chaperonin-10. It’s been found to be detectable as early as 1 week in cows and within 48 hours of conception in women.

If you test for it the old-fashioned way.

The old-fashioned way is call a Rosette Inhibition Test (RIT)—a complicated, cumbersome, and expensive procedure usually done in a lab.

It’s not impossible. You can order a kit. But you would need:

1. Standard microplane reader
2. Precision pipettes and disposable pipette tips
3. 37 degree incubator
4. Centrifuge
5. $550 for 48 strips
6. Expertise

The RIT was also mentioned in“Measurement of EPF for detection of cow pregnancy using rosette inhibition test” in 2008:

“In conclusion, the RIT score for EPF detection can be used as a tool for early detection pregnancy or embryonic death in dairy cows.”² (Laleh, Laleh, Pirany, & Ahrabi, 2008)

EPF seems to work well as an extremely early indicator of conception in several animals and humans.

What exactly is EPF?

“Biochemically, EPF is a homolog of chaperonin 10 and belongs to the heat shock family of proteins with immunosuppressive and growth factor properties [48] and seems involved in the suppression of the maternal response, thereby allowing the continued viability of the early embryo.”³ (Benagiano, Mancuso, Guo, & Di Renzo, 2023)

“The properties of EPF, namely rapid appearance after fertilization, persistence during the first two trimesters of pregnancy and rapid disappearance following the death or surgical removal of the embryo (Morton et al., 1977; Nancarrow et al., 1979) suggests its use as a potential marker for early diagnosis of pregnancy and as a tool to monitor embryonic development. This agrees with the finding of Tinneberg et al. (1985) who found that EPF activities in three patients after interruption fell to control levels within 50 hours.”⁴ (Shahani, Moniz, Chitlange, & Meherji, 1992)

AC Cavanaugh of the University of Queensland published a paper in 1996:

“In this review I document this 20 year quest (undertaken by myself in association with Halle Morton, who made the original discovery of EPF, as well as a number of co-researchers), highlighting the most important milestones and describing recent work”

“…in addition to early pregnancy diagnosis, detection of EPF could be valuable in discriminating between failure to fertilize and failure to implant, as well as in monitoring ‘at risk’ pregnancies.”⁵ (Cavanaugh, 1996)

But despite showing great potential, not much testing has been done for EPF using the Rosette Inhibition Test over the last 50 years.

Throwing Shade

Some recent mentions of EPF reflect skepticism about its usefulness. However, doubts in 2019 about its “future” potential seem to link back to a single source in 1987.

Blackshaw 2019 references Jarvis 2016:

“There are doubts about EPF’s utility for diagnosing early pregnancy (Jarvis 2016) and it has rarely been used since hCG testing became widely available.”⁶ (Blackshaw & Rodger, 2019)

Jarvis 2016 references Chard 1987 (superscript 63 is the endnote pointing to Chard):

“However, there is doubt about the utility of EPF for diagnosing early pregnancy ⁶³ and little has been published on it in the past five years.”⁷ (Jarvis, 2016)

And Chard 1987 can’t be found other than the abstract on Pubmed:

“many workers have been unable to reproduce these findings and the experimental methods have been severely criticized. Substantial further work is needed before a test of this type can be regarded as suitable for routine clinical practice, if indeed it will ever be possible.”⁸ (Chard, 1987)

Jarvis continues in a 2020 article:

“…two putative biological markers, early pregnancy factor (EPF) and embryo-derived platelet activating factor (EDPAF), that had been proposed to be released within 24 hours of fertilisation…[however], since 2001, little work has been published on EPF or EDPAF and any initial promise they may have had for detecting pre-implantation embryos has long since faded.”⁹ (Jarvis, 2020)

There don’t seem to be many articles in between.

You might wonder if a moratorium had been placed on EPF research in conjunction with pushing out the pill.

Determining the fate of preimplantation embryos under natural conditions is arguably better left undone.

But in the presence of CRIPLE, it’s necessary in order to evaluate the true mechanism of action of hormonal and emergency contraception.

Let ‘em Cook

Not everyone has given up on determining the fate of pre-implantation human embryos.

Kahlenborn and Peck are optimistic:

“Since EPFs discovery, almost every study has corroborated its sensitivity and specificity except two (Cooper and Aitken 1981; Thomson et al. 1980). Both of these studies, however, deviated from Morton’s original method of measuring EPF (Smart et al. 1982b). EPF has been described in detail in multiple papers from the 1970s through early 2000. We believe it could be a tool for settling the controversy regarding LNG-EC’s mechanism of action.”

“EPF might also be used to test whether oral contraceptives work at times by causing the destruction of the embryo, as is implied by several papers (Larimore and Stanford 2000; Pierson et al. 2003).”¹⁰ (Kahlenborn, Peck, & Severs, 2015)

They even describe the challenges faced with using EPF, and the blueprint required to overcome them:

“…using EPF to measure abortion rates with LNG-EC would be technically challenging. One would have to follow Morton’s original protocol, which would require time and a sophisticated laboratory. One would have to confirm EPF’s sensitivity and specificity for that particular study by simultaneously studying a sidecontrol group containing early pregnancy and non-pregnancy cohorts. One would have to measure EPF several times at one-day intervals starting with the second day after fertilization to avoid missing very early pregnancies that might expire before having the opportunity to test for them. Finally, one would have to measure EPF when LNG-EC is given both prior to ovulation and after ovulation.” (Kahlenborn, Peck, & Severs, 2015)

It’s been 50 years since the discovery of EPF.

The COVID vaccine was developed in under 12 months.

Developing a test to better understand early embryo loss and pregnancy processes is important for the species. It’s not a matter of science, but of the will.

And if not for the women, do it for the cows.

1. Ghaffari Laleh, V., Ghaffari Laleh, R., Pirany, N., & Moghadaszadeh Ahrabi, M. (2008). Measurement of EPF for detection of cow pregnancy using rosette inhibition test. Theriogenology, 70(1), 105–107. https://doi.org/10.1016/j.theriogenology.2008.02.003 ↩︎
2. Ghaffari Laleh, V., Ghaffari Laleh, R., Pirany, N., & Moghadaszadeh Ahrabi, M. (2008). Measurement of EPF for detection of cow pregnancy using rosette inhibition test. Theriogenology, 70(1), 105–107. https://doi.org/10.1016/j.theriogenology.2008.02.003 ↩︎
3. Benagiano, G., Mancuso, S., Guo, S. W., & Di Renzo, G. C. (2023). Events Leading to the Establishment of Pregnancy and Placental Formation: The Need to Fine-Tune the Nomenclature on Pregnancy and Gestation. International journal of molecular sciences, 24(20), 15420. https://doi.org/10.3390/ijms242015420 ↩︎
4. Shahani, S. K., Moniz, C., Chitlange, S., & Meherji, P. (1992). Early pregnancy factor (EPF) as a marker for the diagnosis of subclinical embryonic loss. Experimental and clinical endocrinology, 99(3), 123–128. https://doi.org/10.1055/s-0029-1211152 ↩︎
5. Cavanagh, A. (1996). Identification of early pregnancy factor as chaperonin 10: implications for understanding its role. Reviews of Reproduction, 1(1), 28-32. Retrieved Jan 27, 2025, from https://doi.org/10.1530/revreprod/1.1.28 ↩︎
6. Blackshaw, B. P., & Rodger, D. (2019). The Problem of Spontaneous Abortion: Is the Pro-Life Position Morally Monstrous?. The New bioethics : a multidisciplinary journal of biotechnology and the body, 25(2), 103–120. https://doi.org/10.1080/20502877.2019.1602376 ↩︎
7. Jarvis G. E. (2016). Early embryo mortality in natural human reproduction: What the data say. F1000Research, 5, 2765. https://doi.org/10.12688/f1000research.8937.2 ↩︎
8. Chard, T., & Grudzinskas, J. G. (1987). Early pregnancy factor. Biological research in pregnancy and perinatology, 8(2 2D Half), 53–56. ↩︎
9. Jarvis G. E. (2020). Misjudging early embryo mortality in natural human reproduction. F1000Research, 9, 702. https://doi.org/10.12688/f1000research.22655.1 ↩︎
10. Kahlenborn, C., Peck, R., & Severs, W. B. (2015). Mechanism of action of levonorgestrel emergency contraception. The Linacre quarterly, 82(1), 18–33. https://doi.org/10.1179/2050854914Y.0000000026 ↩︎