Serum anti-Mullerian hormone (AMH) is a unique biomarker that has a critical role in folliculogenesis as well as steroidogenesis within ovaries. Secretion from preantral and early antral follicles renders AMH as the earliest marker to show ovarian reserve decline.

1. AMH -Novel Biomarker in women
Dr Rajesh V Bendre
Chief Pathologist MD(Path), DNB(Path), DPB
Mumbai

2. INTRODUCTION -
Infertility (subfertility) is a social problem, even
in a country like India, with a staggering
population of about 1 billion, affecting 6.9-9.3%
of the women.
With increasing understanding that age alone
cannot accurately predict ovarian response & that
simply using stimulation strategy to maximizing
oocyte yield is not appropriate for all patients- has
lead to an explosion in the literature regarding the
utility of biomarkers to predict ovarian response.
“Ovarian reserve” has traditionally been used to
describe a woman’s reproductive potential
specifically, the number and quality of oocytes
she possesses. However, commonly used ovarian
reserve markers serve as indicators for oocyte
quantity but are poor predictors of oocyte quality.
Loss of Oocyte Quantity with
advancing age

3. Anti-Müllerian hormone – Ovarain Reserve marker
Maximal expression occurs in
preantral and small antral
follicles1,2
Expression disappears in maturing
pre-ovulatory follicles (expression
restricted to GCs of the cumulus)2
1. Laven et al. J Clin Endocrinol Metab 2004; 89: 318–323;
2. Weenen et al. Mol Hum Reprod 2004;10: 77–83;
3. Cook et al. Fertil Steril 2000; 73: 859–861;
4. La Marca et al. Hum Reprod 2004; 19: 2738–2741;
5. La Marca et al. Hum Reprod 2006; 21: 3103–3107
• AMH is expressed in pre-antral and
small antral follicles.AMH is thus a good
indicator of the ovarian antral follicle
pool & ovarian reserve.
• The ovary-specific expression pattern in granulosa cells of growing non selected follicles
makes AMH an ideal marker for the size of the ovarian follicle pool and also a prognostic
factor for fertility potential.

4. • AMH is a dimeric glycoprotein belonging to
the transforming growth factor-beta (TGF-
β) superfamily. It has an inhibitory effect on
primordial follicle recruitment as well as on
the responsiveness of growing follicles to
FSH in ovary.
• Produced by granulosa cells of pre-antral and
small antral follicles
• Physiological function- prevent excessive
follicle recruitment
• Not cycle dependant-can be measured any day
• Less cycle to cycle variation than FSH.
• Not altered after hormonal therapy.
• Not altered even after down regulation with
GNRH agonist.
Potential applications of Antimullerian hormone (AMH)

5. Antimullerian hormone (AMH)- Age specific ranges

6. INTRODUCTION – Ovarian Reserve Markers

7. INTRODUCTION – Ovarian Reserve Markers
Anti-Mullerian hormone (AMH) as a predictive marker in assisted reproductive technology (ART).
2010 ,Human reproduction update

8. Frequency of discordance by age of serum
FSH and AMH during estradiol-confirmed
menstrual cycle days 2–4 in 5354 women
from U.S. fertility centers.
Discordance was defined when one test
was ‘concerning’ and the other test was
‘reassuring’. Cutpoints for AMH were less
than 0.8 ng/ml (concerning) and at least
0.8 ng/ml (reassuring), and for FSH were at
least 10 IU/l (concerning) and less than
10 IU/l (reassuring).
AMH & FSH

9. Broer et al.2013 have compared AFC and AMH and have shown that sensitivity and
specificity for AMH were 82 and 76%, respectively, and 82 and 80%, respectively, for
AFC.
Comparison of the summary estimates and ROC curves for AMH and AFC showed no
statistical difference in this study. Both AMH and AFC are accurate predictors of excessive
response to ovarian hyperstimulation
AMH & AFC

10. INTRODUCTION – Ovarian Reserve Markers

11. Antimullerian hormone (AMH) & IVF

12. Antimullerian hormone (AMH) & IVF

13. Nelson. Biomarkers of ovarian response.
Fertil Steril 2013. The AMH values are in
pmol/L.
Ovarian response categories dictate risk, and treatment strategies are designed to
minimize risk while maximizing oocyte yield within each response category.
Negligible response means that the conventional criteria for triggering (three
follicles more than or equal to 17 mm) is unlikely to be achieved.
Individualization of controlled ovarian
stimulation in IVF using ovarian reserve
markers:from theory to practice. Human
Reproduction Update,pp. 1–17, 2013
Antimullerian hormone (AMH) & AFC

14. Antimullerian hormone (AMH) & IVF

15. Screening for mutations of the follicle stimulating hormone receptor (FSHR) gene, two
polymorphisms were identified:
• located in the extracellular domain at Position 307, occupied by either alanine (Ala) or
threonine (Thr); and
• located in the intracellular domain at Position 680, occupied either by asparagines (Asn) or
serine (Ser).
Both polymorphic sites are within Exon 10 and give rise to two discrete allelic variants of the
FSHR, i.e. Thr307/Asn680 and Ala307/ Ser680.
FSH Receptor Polymorphism & ovarian response

16. AMH & Polycystic Ovarian Syndrome -

17. PCOS – individual follicles produce more AMH
AMH concentrations in size-matched granulosa cells is
Increased by -
Anovulatory PCOS
Ovulatory PCOS
Normal ovaries
x6 the density of pre-antral follicles in PCOS compared with normal ovary. This increased density
& increased individual granulosa cell production often correlating with high levels (x2 to 3) of
serum AMH.
(Webber et al, 2003 Pigny et al, 2003; Pellatt et al, 2007)
• AMH elevation correlates with PCOS severity and is predictive of insulin resistance
and anovulation. AMH concentrations are very high in PCOS. The higher they are,
the greater the ovulatory disturbance. Possibly because of the inhibitory action of
AMH on aromatase & altered E2 production.
X 4-18
Role of Anti-Müllerian hormone in PCOS-
Individual cell dysfunction in PCOS

18. * Role of AMH Cut off for Diagnosis of PCOD
Iliodromiti et al AMH Threshold for Diagnosis of PCOS. J Clin Endocrinol Metab, August
2013, 98(8):3332–3340

19. Key Features Advantages
Antibodies and calibration identical to
AMH Gen II
Consistent & standardized results. Same
numeric value as AMH Gen II
Increased sensitivity & precision at low
end of analytical range. No
complement interference
Improved support of fertility assessment
Calibrators prepared with recombinant
human AMH vs Bovine
Lyophilized calibrator
Improved accuracy of patient results,
long-term
Increased stability
Less technician handling time &
increased ease of use (vs. MIA)
Efficient and cost effective results
Fully automated random access
instrumentation- Access2, Roche
Speed, quality and flexibility
AMH- Beckman Coulter Gen II – Advancing from ELISA TO CLIA
Laboratory Aspects- Antimullerian hormone (AMH)

20. Laboratory Aspects- Antimullerian hormone (AMH)

21. * Variability in AMH results must take
into account following variables for
appropriate interpretation in clinical
care-
*Biological fluctuation within
individuals; especially obesity, smokers
*exposure to medications (such as
contraceptive hormones, repeated IVF ),
certain ovarian surgeries;
*Laboratory-specific AMH values as
each laboratory provides its own value
ranges and calibration.
Maximizing the clinical utility of
AMH testing in women’s health.
Curr Opin Obstet Gynecol 2014,
26:226–236
Laboratory Aspects- Antimullerian hormone (AMH)

22. Laboratory Data- A pilot study done on IVF patients in collaboration with
group of three gynaecologists - 105 samples for validating AMH ranges
used for stratifying female infertile patients & diagnostic cut off correlating
with PCOS.
AMH Values (ng / ml) FSH
values
mIU/mL
Optimal Satisfactory Low fertility Very low High
4.0 - 6.79 2.19 - 4.0 0.3-1.0 1.0-1.5 1.5-2.19 < 0.3 > 6.79 Average
IVF Protocol Antagonist
control
HCG
Agonist
Trigger
Long down-
regulation
Agonist
control
Flare Agonist
Flare
Agonist /
oocyte
donation
Antagonist
control
Agonist
Trigger
21- 25 (8) 4 3 0 0 0 0 1 4.57
25 - 40 (89) 18 25 13 1 10 7 15 7.92
Above 40 (8) 0 0 0 0 0 8 0 21.34

23. Correlation of AMH levels for stratifying patients for IVF protocol & outcomes-
Laboratory Data- A pilot study done on IVF patients in collaboration with group of
three gynaecologists - 105 samples for validating AMH ranges used for stratifying
female infertile patients & diagnostic cut off correlating with PCOS.
AGE GROUP
AMH Values (ng / ml) FSH values
mIU/mL
Optimal Satisfactory Low fertility Very low High Average
Years 4.0 - 6.79 2.19 - 4.0 0.3-1.0 1.0-1.5 1.5-2.19 < 0.3 > 6.79
21-25 (8) 4 3 0 0 0 0 1
4.57
25 - 40 (89) 18 25 13 1 10 7 15
7.92
Above 40 (8) 0 0 0 0 0 8 0
20.34
IVF,
pregnancy
outcome
10 conceived
out of 22
(44%)
12 conceived out
of 28(40%)
8 conceived out of 24 (33%),
5(above1.5), 1(above1), 2(below1)
4
conceived
out of 15
(26%)
3
conceived
out of 16
(18%)

24. Use of AMH for Risk Stratification & selection of IVF protocol
* Current study used 2.19 as cut off for satisfactory fertility, but lowering
the cut off to 1.5 can also be considered
Correlation of AMH levels for PCOS-
*All 16 patients having AMH above 6.79 ng/ml correlated with clinical
and/or biochemical hyperandrogenemia
*12 patients showed polycystic ovarian morphology while 4 patients
showed normal ovaries
Conclusion- AMH can be used for stratifying patients for IVF with cut off
of 2.19 ng/ml for satisfactory fertility and 6.7 ng/ml as cut off correlates as
an adjuvant in diagnosis of PCOS
Laboratory Data- A pilot study done on IVF patients in collaboration with
group of three gynaecologists - 105 samples for validating AMH ranges
used for stratifying female infertile patients & diagnostic cut off correlating
with PCOS.