Follow-up strategies for IVF patients slightly vary among reproductive centers. Most reproductive centers adopt blood pregnancy tests 14 days after embryo transfer [15, 16], while some centers implement hospital pregnancy tests on the 11th or 12th day after the transfer [17, 18]. In our center, HCG demonstrated spikes two to three times at 72–96-h intervals following pregnancy tests. Approximately 30 days after embryo transfer, a viability ultrasound was performed to determine whether a fetal heartbeat existed. If the HCG levels grew poorly, the number of follow-ups prior to the initial B-mode ultrasound should be increased to avoid the risk of ectopic pregnancy and threatened abortion. However, as a matter of fact, women undergoing IVF usually feel anxious due to their own infertility and the unpredictability of pregnancy progress. Such stress leads to several behavioral changes [19]. For instance, some patients start performing pregnancy tests just 5–6 days after embryo transfer, and if the urine β-HCG test is positive, they immediately go to the hospital for a blood test verification.
Stress fluctuates during IVF treatment; the period from oocyte retrieval to embryo transfer and the waiting time for pregnancy tests were specifically linked to high degrees of anxiety and stress [20, 21]. For patients, the stress of waiting is the direct cause of taking a pregnancy test beforehand. In this study, we found that patients whose pregnancy test time was ≤ 9 days from the day of embryo transfer were younger, indicating that younger patients were more eager to undergo a pregnancy test earlier. In contrast, older patients showed varying expectation levels and complied better, probably because of higher previous childbirth ratios, with many of them taking pregnancy tests within the prescribed period. The ≥ 13 days group was not inferior to the other two groups in terms of factors conducive to embryo implantation, such as endometrial thickness, blastocyst transfer rate, and good-quality embryo transfer rate. Thus, we may consider that the reason for late pregnancy test time was linked to the difference in patient compliance. Additionally, despite the earlier implantation requirement following blastocyst transfer, the overall pregnancy time preference of blastocyst transfer patients was later than the D3 embryo transfer patients, suggesting that the pregnancy test time was decided based on patient compliance.
For the general population, patients taking pregnancy tests on days 9 or 10 after transfer exhibited the highest clinical pregnancy rate, low early miscarriage rate, and high live birth rate. Even after women aged over 35 years and those with previous childbirth(s) were excluded, the population taking pregnancy test on day 9–10 after transfer exhibited a lower early miscarriage rate and higher live birth rate. The too-early testing behavior did not necessarily predict a favorable pregnancy outcome. The reasons are stated as follows: 1. Some patients are detected positive for urine HCG in an extremely early period after transfer, suggesting that the embryo cleaves rapidly, with an advanced implantation compared with an embryo developing at a normal rate. The further development potential of rapidly-cleaving embryo remains controversial, and some scholars suggest that the rapidly-cleaving embryo is associated with a markedly increased chromosomal abnormality rate. It is interesting that the fast cleaving rate is related to similar chromosomal patterns to the slowly-cleaving counterparts. Embryos that exhibit 9 blastomeres after insemination for 62 h may experience chromosomal abnormality at an identical probability to that of their slowly-cleaving counterparts. Compared to embryos that contain 8 cells, those containing 9–11 cells have a lower development potential [22]. 2. As suggested in some articles, stress level in the IVF process affects reproductive outcome [23]. An early pregnancy test time demonstrates that the patient is anxious to some extent, and when she obtains the positive result, she may be worry about the blood HCG level. As confirmed by a study concerning the association of emotion with early miscarriage, emotional health deficiency increases the risk of miscarriage [24]. The reasons for poor pregnancy outcome at a late pregnancy test time may be due to the following reasons. Uterine receptivity occurs around 5–7 days after ovulation and lasts for 4 days, when embryo implantation commences. To ensure a successful implantation, the growth and maintenance of blood vessels should be coordinated at the maternal-embryo interface, so that the environment offered is nourished [25]. Patients with a pregnancy test time around 9–10 days are in the stage of embryonic post invasion and persistent uterine decidualization [26–28]. If these patients come to the hospital for a blood pregnancy test during this period, they can receive individualized luteal phase support (LPS) and other medications (e.g. low-molecular-weight heparin or glucocorticoid adjustments) based on the results of hormone levels, D2 dimers, immunoassays, and other tests, which may better facilitate endometrium remodeling, including an influx of uterine-specific NK cells and the exclusion of maternal B and T cells [27]. This is beneficial for continuous embryo implantation. However, if the test time around 9th-10th day is missed, the patient may miss the optimal luteal support adjustment time, leading to a poor pregnancy outcome.
There were no differences in the late-term miscarriage rate or the gestational week of delivery among the three groups, suggesting that women with a relatively stable early pregnancy showed no differences in pregnancy outcomes during later gestation. The stability of early pregnancy under identical blastocyst transfer rate and good-quality transfer ratio, i.e., under equal development potentials of embryos, can be improved in the following ways: 1. Strengthening the post-transfer lifestyle and psychological support. Some treatments or therapies, including relaxation training, medical clowning, coping cards, and hypnosis during embryo transfer, can be used to relieve the anxiety and stress caused by the uncertainties of pregnancy outcomes [29–32]. We should not object to the patients performing an early pregnancy test at home but help them in properly accepting the self-tested urine HCG results. 2. Progesterone administration remains controversial regarding dosages, routes, and timing [33–35]. In our reproductive center, a combination therapy with varying routes is used. For patients with deficient serum progesterone levels under a single treatment, our therapy seems a plausible option for maintaining adequate progesterone exposure. Besides, it can eliminate the increase in early miscarriage rates resulting from improper medications, drug allergies, and incompatible luteal support. There was no difference in the number of women with natural cycle FET among the three groups, and the luteal support protocols were consistent for each procedure of fresh/frozen embryo transfer. According to some literature, the serum progesterone measurements on the day of pregnancy test are linked to the ongoing pregnancy rate and live birth ratio [36–38]. In our study, the progesterone levels on the day of pregnancy test did not differ among the three groups. Nevertheless, Manuel Alvarez et al. argued that the appropriate time point for initiating an individualized LPS should be the day prior to blastocyst transfer [39]. Further prospective research is required to determine whether progesterone detection one week following the transfer and the corresponding adjustment of luteal support protocol is necessary.
This study has some shortcomings, regarding the fact that there are no data on pregnancy tests prior to the test at the hospital (self tests), and this is a factor that interferes with the final conclusion, which is a limitation of retrospective analysis. The cause analysis of early miscarriage (especially the embryo karyotype abnormalities, which are the most frequent cause of miscarriage during the first trimester) is lacking in the present study because not every miscarriage patient can accept the embryo chromosomal analysis. Replication and extension of our study can be a valuable direction for future prospective research.