By Rebecca H. Allen, MD, MPH, Editor

SYNOPSIS: In this prospective cohort pilot study, the authors validated a flow cytometry protocol for detecting fetal red blood cells and determined that fetal red blood cell exposure in first-trimester uterine aspiration was well below the calculated threshold for maternal Rh sensitization.

SOURCE: Horvath S, Tsao P, Huang ZY, et al. The concentration of fetal red blood cells in first-trimester pregnant women undergoing uterine aspiration is below the calculated threshold for Rh sensitization. Contraception 2020; March 3. doi: 10.1016/j.contraception.2020.02.011. [Online ahead of print].

The authors of this study developed a flow cytometry protocol to detect fetal red blood cells (RBCs) that relied on dual staining with hemoglobin F and carbonic anhydrase to distinguish fetal from adult RBCs, and then further distinguished maternal F cells from fetal RBCs by the brightness of staining for hemoglobin F. This flow cytometry protocol then was evaluated with an in vivo pilot study of 42 participants. Based on previous studies, the authors considered 0.1 mL of RBCs the minimum volume of fetomaternal hemorrhage capable of causing Rh sensitization. The average pregnant woman’s blood volume in early pregnancy is 4,000 mL. Therefore, the target fetal RBC concentration for detection was 0.1/4,000 mL or 250 fetal RBCs per 10 million total adult RBCs. Women undergoing uterine aspiration for spontaneous or induced abortion less than 12 completed weeks gestation were recruited. Women with hemoglobinopathies or vaginal bleeding prior to enrollment were excluded. Blood was drawn before and after uterine aspiration and samples were run within 72 hours of collection. The authors were able to validate their flow cytometry protocol and then apply it to the study subjects. A total of 37 of 42 participants (88%) had pre- and post-aspiration samples sufficient for analysis. All fetal RBC samples were below the calculated threshold for Rh sensitization of 250 fetal RBCs per 10 million adult RBCs. The fetal RBC concentration increased from pre-aspiration (mean 4.5, median 0, range 0-57) to post-aspiration (mean 8.6, median 2, range 0-32). There was no statistically significant difference in fetal RBC concentration by gestational age, use of sharp curettage, pregnancy history, or participant demographics.

COMMENTARY

The need for anti-D immunoglobulin (RhoGAM) after first-trimester events, such as threatened abortion, spontaneous abortion, molar pregnancy, and induced abortion, lacks consensus.^1,2

International recommendations differ, for example, with the Royal College of Obstetricians and Gynaecologists not requiring RhoGAM for abortion less than 12 weeks unless the uterus is surgically evacuated, while the American College of Obstetricians and Gynecologists (ACOG) states that administering a 50 mcg dose in early pregnancy loss “should be considered.”^1-4 However, not every facility has the 50 mcg dose available for first-trimester patients and many administer the full 300 mcg dose, which may be a waste of resources. Currently, no synthetic anti-D immunoglobulin is available, and our supply comes from human donors. Since approximately 15% of women in the United States are Rh negative, this recommendation affects new patients, as well as emergency departments, hospitals, and abortion clinics. Therefore, it is laudable that the authors completed this pilot study.

Traditional measurement of fetomaternal hemorrhage with Kleihauer-Betke (K-B) testing is limited because the lower limit of detection is 4,000 fetal RBCs per 10 million adult RBCs. In addition, the K-B test does not distinguish between fetal RBCs and maternal F cells, which can lead to false-positive results. There-fore, the authors opted to use a flow cytometry protocol, which is more sensitive, and then validated it by looking at both maternal F cells and fetal RBCs. Previous studies provided the authors with a plausible threshold concentration for fetomaternal hemorrhage during the first trimester (250 fetal RBCs per 10 million total adult RBCs). The fact that the authors did not find any subject approaching this threshold is encouraging. Luckily, the authors will be pursuing this work with a larger sample of patients; therefore, more data will be accrued.

The National Abortion Federation already changed its guideline in March 2019 to allow for waiving Rh testing and RhoGAM administration for Rh-negative patients undergoing surgical abortion up to eight weeks gestation and medication abortion up to 10 weeks gestation, based on emerging evidence.^2,5,6 This change may allow women seeking induced abortion to pay less out of pocket for the procedure. The change also facilitates medication abortion via telemedicine, which is becoming more important in the COVID-19 pandemic.⁷ I suspect many other U.S. providers likely will wait for ACOG to change its recommendation before changing practice in the first trimester. At the very least, I wish more hospitals would stock the 50 mcg dose of RhoGAM to conserve supplies.

REFERENCES

1. Sperling JD, Dahlke JD, Sutton D, et al. Prevention of RhD alloimmunization: A comparison of four national guidelines. Am J Perinatol 2018;35:110-119.
2. Wiebe ER, Campbell M, Aiken ARA, Albert A. Can we safely stop testing for Rh status and immunizing Rh-negative women having early abortions? A comparison of Rh alloimmunization in Canada and the Netherlands. Contraception: X 2019;1. https://doi.org/10.1016/j.conx.2018.100001
3. [No authors listed]. ACOG Practice Bulletin No. 200: Early pregnancy loss. Obstet Gynecol 2018;132:e197-e207.
4. Committee on Practice Bulletins-Obstetrics. ACOG Practice Bulletin No. 181: Prevention of Rh D alloimmunization. Obtset Gynecol 2017;130:e57-70.
5. Hollenbach SJ, Cochran M, Harrington A. “Provoked” feto-maternal hemorrhage may represent insensible cell exchange in pregnancies from 6 to 22 weeks gestational age. Contraception 2019;100:142-146.
6. Mark A, Foster AM, Grossman D, et al. Foregoing Rh testing and anti-D immunoglobulin for women presenting for early abortion: A recommendation from the National Abortion Federation's Clinical Policies Committee. Contraception 2019;99:265-266.
7. Raymond E, Grossman D, Mark A, et al. Medication abortion: A sample protocol for increasing access during a pandemic and beyond. UC Davis. 2020. https://escholarship.org/uc/item/02v2t0n9