Emergency contraception in the emergency department.

BACKGROUND: On June 24, 2022, the Supreme Court overturned Roe v. Wade, which will limit legal abortion in many areas of the U.S., making the need for effective emergency contraception even more critical. METHODS: This narrative review focuses on the approach to providing safe and effective emergency contraception in the emergency department (ED) with a focus on agents that are used in the U. S.; however, many of the agents discussed are also available and utilized in other countries. RESULTS: Emergency contraception methods included in this review are, ulipristal, levonorgestrel, combined oral contraceptive pills, and copper intrauterine devices (IUDs). CONCLUSION: The efficacy of products used for emergency contraception depend on patient and temporal factors. Emergency physicians must have an understanding of the optimal use of these agents to prevent unwanted pregnancy, particularly in the setting of restricted abortion access.

A national analysis of ED presentations for early pregnancy and complications: Implications for post-Roe America.

BACKGROUND: Most obstetric emergencies are initially managed in the emergency department (ED). The Supreme Court decision of Dobbs v. Jackson Women's Health Organization, overturning Roe v. Wade, in June 2022, eliminated constitutional protection of abortion rights, allowing states to swiftly enact laws that can greatly change reproductive medicine. In this post-Roe landscape, the ambiguity and uncertainty being imposed on clinicians regarding the legality of certain interventions may have catastrophic effects. To understand and plan for the changes that will come and attempt to mitigate adverse outcomes, the authors first assessed the current state of pregnancy-related complication care in the ED setting. This study utilized data obtained from the National Hospital Ambulatory Medical Care Survey (NHAMCS) to evaluate trends in pregnancy-related ED visits from 2016 to 2020 that could be impacted by restricted abortion access and trigger laws. The authors subsequently analyzed the legislative changes and translated the pertinent ones to dispel misunderstandings and provide a framework for appropriate medical practice. METHODS: The retrospective study utilized data from the NHAMCS database from 2016 to 2020, encompassing an estimated total of 4,556,778 pregnancy-related ED visits. NHAMCS is a multi-stage probabilistic sample collected by the National Center for Health Statistics (NCHS) at the Centers for Disease Control and Prevention (CDC) using an annual survey of EDs in the United States. All data were summarized using descriptive statistics such as proportions and 95% confidence intervals Furthermore, the supreme court decision was analyzed in addition to multiple state laws and legal texts. The findings were summarized and discussed. RESULTS: The majority (79.4%) of all studied visits were for patients between the ages of 18-34 years, capturing those in peak reproductive years. This age group also comprised of the bulk (76.4%) of visits for pathologic pregnancies, including ectopic and molar pregnancies, and 79.8% of visits for a spontaneous miscarriage or threatened miscarriage in early pregnancy. Black patients accounted for 25.7%, white patients 70.1%. Regarding ethnicity, patients were separated into Hispanic and non-Hispanic, with Hispanic patients comprising 27% of all ED visits for included diagnoses between 2016 and 2020. Most visits for complications following an induced abortion occurred in the south (70.8%) and were nearly twice as likely to occur in non-metropolitan areas. Approximately 18% patients presenting with a pathologic pregnancy required admission to the hospital and approximately 50% of those visits for pathologic pregnancies and visits for bleeding in pregnancy had a procedure in the ED (49.8% and 49.5%). There were 111,264 estimated visits in which methotrexate was administered, amounting to approximately 1 in 7 visits for ectopic or molar pregnancy. In this data set, approximately 14,000 miscarriage and early bleeding patients received misoprostol. CONCLUSION: Pregnancy-related ED visits comprise of a significant proportion of emergency care. As it relates to many of the trends previously elucidated on, the true extent of the burden cannot be predicted. It must be emphasized that contrary to popular belief, Dobbs v. Jackson does not prohibit termination of pregnancy in the setting of life-threatening conditions to the mother, including ectopic pregnancy, preeclampsia, and others, but the resultant uncertainty and ambiguity surrounding the constitutional change is leading to an over-compliance of the law, necessarily obstructing reproductive health care. The authors recommend that physicians be mindful of the rapidly-evolving laws in their particular state, and to also practice in accordance with Emergency Medical Treatment and Active Labor Act (EMTALA). Patient safety must be prioritized.

Understanding Psychosocial Determinants of PrEP Uptake Among Cisgender Women Experiencing Heightened HIV Risk: Implications for Multi-Level Communication Intervention.

Pre-exposure prophylaxis (PrEP) is a highly effective daily pill that decreases the likelihood of HIV acquisition by up to 92% among individuals at risk for HIV. PrEP can be discretely used, autonomously controlled, and in place at the time of risk exposure, making it an especially promising method for HIV prevention for cisgender women (CGW). But, PrEP is underutilized by CGW relative to the demonstrable need. We apply the Integrative Model of Behavioral Prediction to identify the critical psychosocial factors that shape CGW's intentions to use PrEP and their relevant underlying beliefs. We surveyed (N = 294) community- and clinic-recruited PrEP eligible CGW to understand the relative importance of attitudes, norms, and efficacy in shaping PrEP intentions. We utilized structural equation modeling to identify the relevant paths. We inspected the summary statistics in relation to three message three selection criteria. We identified beliefs that demonstrated (1) an association with intention, (2) substantial room to move the population, (3) practicality as a target for change through communication intervention. Results show that PrEP awareness was low. When women learned about PrEP, they voiced positive intentions to use it. There were significant and positive direct effects of SE (0.316***), attitudes (0.201**), and subjective norms (0.249***) on intention to initiate PrEP. We illustrate the strategic identification of beliefs within the relevant paths using the 3 belief selection criteria. We also discuss implications for social and structural communication interventions to support women's HIV prevention.

Factors Associated with Intention to Initiate Pre-exposure Prophylaxis in Cisgender Women at High Behavioral Risk for HIV in Washington, D.C.

Pre-exposure prophylaxis (PrEP) for HIV prevention is underutilized by cisgender women at risk for HIV in the USA. Published research on PrEP initiation among cisgender women at risk for HIV focuses on identifying barriers and facilitators associated with intention to initiate, but few apply a behavioral theoretical lens to understand the relative importance of these diverse factors. This study provides a theoretically grounded view of the relative importance of factors associated with intention to initiate PrEP. We conducted an anonymous, cross-sectional survey of 1437 cisgender women seeking care at family planning and sexual health clinics to evaluate hypothesized barriers and facilitators of PrEP initiation. We categorized cisgender women with ≥ 3 behavioral risk-factors as "high-risk" for HIV acquisition; 26.9% (N = 387) met high-risk criterion. Among cisgender women in the high-risk sample, the majority were Black and single. Perceived risk of HIV acquisition was low and 13.7% reported intention to initiate PrEP. Positive attitudes toward PrEP, self-efficacy, perceived support from medical providers and social networks, and prior discussion about PrEP with medical providers were associated with intention to initiate PrEP; stigma was negatively associated. Background characteristics (other than age), risk factors for HIV acquisition, prior awareness of PrEP, and perceived risk of HIV were not associated with uptake intention. These findings support interventions that center on the role of providers in the provision of PrEP and on social networks in destigmatization of PrEP use.

Toxicity of abortifacients: A review for physicians in the post roe era.

BACKGROUND: On June 24, 2022, the Supreme Court overturned Roe v. Wade, which will limit legal abortion in many areas of the U.S. Over half of abortions in the U.S. are performed using medication as opposed to surgical techniques. With widespread access to agents that are used for medication abortion, there may be an increase in emergency department presentations related to improper or unsupervised use of these medications. METHODS: This narrative review focuses on the contraindications, adverse effects, and toxicities of the most common agents used for medication abortion in the U.S. RESULTS: Medications included in this review are mifepristone, misoprostol, and methotrexate. Each of these medications has a unique adverse effect and toxicity profile. CONCLUSION: Agents used for medication abortion have unique contraindications and adverse effects. Improper or unsupervised use may occur in the setting of limited abortion access and emergency medicine physicians are on the front lines in managing these presentations.

Age-related factors associated with intention to initiate pre-exposure prophylaxis among cisgender women in Washington D.C.

Background: Pre-exposure prophylaxis (PrEP) utilization among cisgender women (subsequently 'women') is low across age groups, relative to their risk of HIV acquisition. We hypothesize that age-related differences in psychosocial factors also influence women's intention to initiate oral PrEP in Washington, D.C. Methods: A secondary analysis of a cross-sectional survey data was performed to evaluate factors influencing intention to initiate oral PrEP among women seen at a family planning and a sexual health clinic. A bivariate analysis was performed to identify differences by age group in demographic characteristics, indications for PrEP, and attitudes toward PrEP; we then performed additional bivariate analysis to assess these variables in relation to PrEP intention. Results: Across age groups, perceived risk of HIV acquisition was not significantly different and was not associated with intention to initiate PrEP. Awareness of and attitude toward PrEP, injunctive norms, descriptive norms, and self-efficacy were not different across age, however there were significant age-associated differences in relation to PrEP intention. Specifically, among 18-24-year-olds, intention to start PrEP was associated with support from provider (p = 0.03), main sexual partner (p < 0.01), and peers (p < 0.01). For women 25-34 years old, having multiple sexual partners (p = 0.03) and support from casual sexual partners (p = 0.03) was also important. Among women 35-44 years old, prior awareness of PrEP (p = 0.02) and their children's support of PrEP uptake (p < 0.01) were associated with intention to initiate PrEP. Among 45-55 year-old women intention to initiate PrEP was positively associated with engaging in casual sex (p = 0.03) and negatively associated with stigma (p < 0.01). Conclusion: Overall, there were more similarities than differences in factors influencing intention to initiate PrEP across age groups. Observed differences offer an opportunity to tailor PrEP delivery and HIV prevention interventions to increase awareness and uptake for cisgender women.

Research showing how women's age affects their decision to start pre-exposure prophylaxis to prevent HIV in Washington, DC We know that many women need pre-exposure prophylaxis (PrEP) to prevent HIV infection in Washington, DC, yet the number of women who use PrEP is lower than expected. The study was done to determine whether a woman's age affects her decision to start PrEP. This is important because providers need to know the factors affecting a woman's decision to start PrEP in order to increase PrEP use in at-risk patients. We re-analyzed a set of data that was generated from surveys given to women in a women's health clinic. The surveys asked women about their background, including risk factors for HIV infection, and their awareness of and attitudes toward PrEP. We also asked women questions that were meant to determine their belief in their own ability to make decisions as well as the degree to which other people in their life affect their own decisions. We wanted to know whether age affects the degree to which these characteristics affect women's decision to start PrEP. Overall, there were more similarities than differences between age groups when looking at how different factors affect the decision to start PrEP. Across age groups, there were no differences in how women view their risk of HIV infection. We found that 18-24-year-olds were more likely to start PrEP when they felt support from their provider, main sexual partner, and peers. 25-34-year-olds were more likely to start PrEP when they felt this same source of support and had had multiple sexual partners. 35-44-year-olds were more likely to start PREP when they already had awareness of PrEP or had their children's support of their PrEP use. 45-55-year-olds were more likely to start PrEP when they had engaged in casual sex and were less likely to start PrEP if they expected negative judgement for it. Even though there were significant similarities between age groups, the differences that we found offer an opportunity to tailor PrEP awareness and interventions to promote PrEP use among women.

Decision and economic analysis of hostile abortion laws compared with supportive abortion laws.

BACKGROUND: On June 24, 2022, the US Supreme Court overturned Roe v Wade in Dobbs v Jackson Women's Health Organization. Therefore, several states banned abortion, and other states are considering more hostile abortion laws. OBJECTIVE: This study aimed to assess the incidence of adverse maternal and neonatal outcomes in the hypothetical cohort where all states have hostile abortion laws compared with the pre-Dobbs v Jackson cohort (supportive abortion laws cohort) and examine the cost-effectiveness of these policies. STUDY DESIGN: This study developed a decision and economic analysis model comparing the hostile abortion laws cohort with the supportive abortion laws cohort in a sample of 5.3 million pregnancies. Cost (inflated to 2022 US dollars) estimates were from a healthcare provider's perspective, including immediate and long-term costs. The time horizon was set to a lifetime. Probabilities, costs, and utilities were derived from the literature. The cost-effectiveness threshold was set to be at $100,000 per quality-adjusted life year. Probabilistic sensitivity analyses using the Monte Carlo simulation with 10,000 simulations were performed to assess the robustness of our results. The primary outcomes included maternal mortality and an incremental cost-effectiveness ratio. The secondary outcomes included hysterectomy, cesarean delivery, hospital readmission, neonatal intensive care unit admission, neonatal mortality, profound neurodevelopmental disability, and incremental cost and effectiveness. RESULTS: In the base case analysis, the hostile abortion laws cohort had 12,911 more maternal mortalities, 7518 more hysterectomies, 234,376 more cesarean deliveries, 102,712 more hospital readmissions, 83,911 more neonatal intensive care unit admissions, 3311 more neonatal mortalities, and 904 more cases of profound neurodevelopmental disability than the supportive abortion laws cohort. The hostile abortion laws cohort was associated with more cost ($109.8 billion [hostile abortion laws cohort] vs $75.6 billion [supportive abortion laws cohort]) and 120,749,900 fewer quality-adjusted life years with an incremental cost-effectiveness ratio of negative $140,687.60 than the supportive abortion laws cohort. Probabilistic sensitivity analyses suggested that the chance of the supportive abortion laws cohort being the preferred strategy was more than 95%. CONCLUSION: When states consider enacting hostile abortion laws, legislators should consider an increase in the incidence of adverse maternal and neonatal outcomes.

Gabapentin as an adjunct for pain management during dilation and evacuation: A double-blind randomized controlled trial.

OBJECTIVES: To assess the analgesic efficacy of preoperative gabapentin among patients undergoing dilation and evacuation (D&E) with moderate sedation. STUDY DESIGN: We conducted a randomized, controlled, double-blind trial among patients undergoing same-day D&E at 14 to 19 weeks gestation under moderate sedation. We randomized participants 1:1 to gabapentin 600 mg or placebo after cervical preparation at least 1 hour prior to D&E. We assessed pain using a 100-mm visual analog scale before, during, and after the procedure. The primary outcome was postoperative recall of maximum procedural pain with a 13-mm a priori threshold for clinical significance. We standardized initial fentanyl and midazolam dosing. We assessed satisfaction with pain control, nausea, and vomiting via Likert scales and anxiety using a validated instrument. RESULTS: We enrolled 126 participants and randomized 61 to gabapentin and 65 to placebo, with study medication administered a mean of 211 (SD 64) minutes preoperatively. Recall of maximum pain was 41 mm for gabapentin and 49 mm for placebo (p = 0.24). Gabapentin resulted in reduced pain during uterine aspiration (56 vs 71 mm, p= 0.003) compared to placebo, but not for any other time points. The gabapentin group had higher satisfaction (78% vs 65% very or somewhat satisfied, p= 0.01). Median fentanyl dose was lower in the gabapentin group (75 vs 100 mcg, p = 0.005). Midazolam dose, nausea, vomiting, and anxiety did not differ between groups. No serious adverse events occurred in the gabapentin group. Sedation reversal was not required. CONCLUSIONS: The addition of gabapentin to moderate sedation during D&E did not result in lower maximum recalled procedural pain. Gabapentin resulted in reduced intra-operative pain during uterine aspiration and increased satisfaction with pain control. IMPLICATIONS: Gabapentin reduces intraoperative pain and improves satisfaction with pain management when administered prior to second-trimester surgical abortion and may be considered as an adjunct to intravenous sedation. Moderate sedation may impair assessment and recall of pain. Additional research is needed to identify the most effective pain management regimens for D&E.

Intention to Initiate HIV Pre-exposure Prophylaxis Among Cisgender Women in a High HIV Prevalence U.S. City.

OBJECTIVE: Our objective was to identify the individual, interpersonal, community, health-system, and structural factors that influence HIV pre-exposure prophylaxis (PrEP) initiation among cisgender women seeking sexual and reproductive health care in a high HIV prevalence community to inform future clinic-based PrEP interventions. METHODS: We collected anonymous, tablet-based questionnaires from a convenience sample of cisgender women in family planning and sexual health clinics in the District of Columbia. The survey used the lens of the socio-ecological model to measure individual, interpersonal, community, institutional, and structural factors surrounding intention to initiate PrEP. The survey queried demographics, behavioral exposure to HIV, perceived risk of HIV acquisition, a priori awareness of PrEP, intention to initiate PrEP, and factors influencing intention to initiate PrEP. RESULTS: A total of 1437 cisgender women completed the survey. By socio-ecological level, intention to initiate PrEP was associated with positive attitudes toward PrEP (odds ratio [OR], 1.56; 95% confidence interval [CI], 1.13-2.15) and higher self-efficacy (OR, 1.32; 95% CI, 1.02-1.72) on the individual level, perceived future utilization of PrEP among peers and low fear of shame/stigma (OR, 1.65; 95% CI, 1.33-2.04) on the community level, and having discussed PrEP with a provider (OR. 2.39; 95% CI, 1.20-4.75) on the institutional level. CONCLUSION: Our findings highlight the importance of multilevel, clinic-based interventions for cisgender women, which promote sex-positive and preventive PrEP messaging, peer navigation to destigmatize PrEP, and education and support for women's health medical providers in the provision of PrEP services for cisgender women.

Integrating HIV Pre-Exposure Prophylaxis into Family Planning Care: A RE-AIM Framework Evaluation.

We aimed to systematically evaluate the feasibility of integrating HIV prevention services, including pre-exposure prophylaxis (PrEP), into a family planning setting in a high-prevalence community. We used the RE-AIM Framework (Reach, Efficacy, Adoption, Implementation, Maintenance) to evaluate the integration of HIV prevention services into a family planning clinic over 6 months. Before the integration, PrEP was not offered. We implemented a staff training program on HIV PrEP. We determined the proportion of women presenting to the clinic who were screened, eligible for, and initiated PrEP through chart review. We assessed staff comfort with PrEP pre- and post-integration. We compared planned and actual implementation, interviewed staff to determine barriers and facilitators, and tracked systems adaptations. We assessed maintenance of PrEP after the study concluded. There were 640 clinical encounters for 515 patients; the rate of HIV counseling and PrEP screening was 50%. The rate was 10% in month 1 and peaked to 65% in month 3. Nearly all screened patients were eligible for PrEP (98.4%) and 15 patients (6%) initiated PrEP. Staff knowledge and comfort discussing PrEP improved after education. Facilitators included partnering with local experts, continuing education, clinical tools for providers, and patient education materials. Barriers included competing priorities during clinical encounters, limited woman-centered patient education materials, and insurance-related barriers. Embedding HIV prevention services in the family planning setting was feasible in this pilot. The proportion of women screened for PrEP rapidly increased. In this high HIV prevalence community, nearly all screened women were eligible and 6% initiated PrEP.

A factorial-design randomized controlled trial comparing misoprostol alone to Dilapan with misoprostol and comparing buccal to vaginal misoprostol for same-day cervical preparation prior to dilation & evacuation at 14 weeks 0 days-19 weeks 6 days gestation.

OBJECTIVES: To compare procedure times following same-day cervical preparation using misoprostol 400 mcg alone or misoprostol 400 mcg plus hygroscopic dilators for dilation and evacuation (D&E) before 20 weeks gestation and to compare side effects of buccal and vaginal misoprostol administration. STUDY DESIGN: We randomized women undergoing D&E at 14 weeks 0 days-19 weeks 6 days gestation to receive (1) hygroscopic dilators or not and (2) buccal or vaginal misoprostol using a 2 × 2 factorial design. We assessed two primary outcomes: (1) total procedure time, defined as time to insert hygroscopic dilators plus D&E time, and (2) side effects of misoprostol 4-6 h after initiation of cervical preparation using a 5-point Likert scale assessing nausea, emesis, diarrhea, chills and cramps. RESULTS: We randomized 163 women and 161 completed the study. We completed all procedures in one day. Mean total procedure time was 14.0 and 10.8 min. with and without hygroscopic dilators (difference 3.2 minutes, 95% CI 1.7, 4.6). Mean D&E procedure time was 0.7 (95% CI -0.8, 2.1) min longer without hygroscopic dilators. Initial cervical dilation was 15.6 and 11.7 mm with and without hygroscopic dilators (difference 3.9 mm, 95% CI 3.1, 4.8). Participants receiving buccal misoprostol reported less chills (1.9) than women receiving vaginal misoprostol (2.3), p = 0.04. CONCLUSIONS: Hygroscopic dilators with misoprostol requires more time and increases cervical dilation without shortening D&E time when used for cervical preparation 4-6 h prior to D&E before 20 weeks. Women receiving vaginal misoprostol may have more chills compared to buccal misoprostol. IMPLICATIONS: Adding hygroscopic dilators to misoprostol for same day D&E procedures at less than 20 weeks gestation increases total intervention time without reducing D&E time and is less favored by patients. Clinical judgment requires balancing relative effectiveness with patient preference. Further studies should evaluate the side effect profile of vaginal misoprostol.

A randomized controlled trial evaluating same-day mifepristone and misoprostol compared to misoprostol alone for cervical preparation prior to second-trimester surgical abortion.

OBJECTIVE: We evaluated initial cervical dilation with the addition of oral mifepristone to vaginal misoprostol as cervical preparation for same-daysecond-trimester dilation and evacuation (D&E). STUDY DESIGN: Women desiring abortion between gestational ages 14weeks 0days and 19weeks 6days were randomized to 200-mg mifepristone or identical placebo immediately followed by 400-mcg misoprostol vaginally 4-6h prior to D&E. Primary outcome was cervical dilation assessed by largest Hegar dilator passed without resistance. Secondary outcomes included total procedure time and participant and provider perceptions. We had 90% power to detect a 2-mm change in initial cervical dilation with a mean of 10mm (SD=3.0mm), requiring 48 participants in each arm. RESULTS: Of 100 women enrolled, 96 were randomized and completed the study. Age, race, gestational age (mean 17.4weeks, SD=1.3) and parity did not significantly differ. Mean initial Hegar dilation measurements were 11.7 and 10.9mm in the mifepristone and placebo groups, respectively, with difference of 0.8 [95% CI=-0.4, 2.0mm]. We found total procedure times of 11.8 and 13.0min, respectively (difference of 1.2min [95% CI=-2.4, 4.8min]. Participant and provider perceptions did not differ. All 96 procedures were completed without hemorrhage, cervical laceration or other observed complications. CONCLUSION: The addition of mifepristone to vaginal misoprostol did not provide a significant increase in cervical dilation compared to misoprostol alone as cervical preparation 4-6h prior to D&E at 14weeks through 19weeks 6days. IMPLICATIONS: Adding mifepristone for a short interval (4-6h) did not improve cervical preparation with misoprostol prior to D&E at 14-19weeks. Future studies should evaluate alternative timing intervals of medications for this purpose.

Use of cell-free DNA in the investigation of intrauterine fetal demise and miscarriage.

OBJECTIVE: To estimate whether cell-free DNA is present in nonviable pregnancies and thus can be used in diagnostic evaluation in this setting. METHODS: We conducted a prospective cohort study of 50 participants at MedStar Washington Hospital Center, Washington, DC, between June 2013 and January 2014. Included were women with pregnancies complicated by missed abortion or fetal demise. All gestational ages were considered for study participation. Participants with fetal demise were offered the standard workup for fetal death per the American College of Obstetricians and Gynecologists. Maternal blood samples were processed to determine the presence of cell-free DNA, the corresponding fetal fractions, and genetic abnormalities. RESULTS: Fifty samples from nonviable pregnancies were analyzed. The average clinical gestational age was 16.9 weeks (standard deviation 9.2). The mean maternal body mass index was 30.3 (standard deviation 9.1). Seventy-six percent (38/50) of samples yielded cell-free DNA results, that is, had fetal fractions within the detectable range of 3.7-65%. Among the 38, 76% (29) were classified as euploid, 21% (8) as trisomies, and 3% (1) as microdeletion. A cell-free DNA result was obtained more frequently at ultrasonographic gestational ages of 8 weeks or greater compared with less than 8 weeks (87.9% [n=29/33, 95% confidence interval (CI) 72.7-95.2; and 52.9%, n=9/17, 95% CI 31.0-73.8] of the time, respectively, P=.012). Time from demise was not associated with obtaining a result. CONCLUSION: Among nonviable pregnancies, cell-free DNA is present in the maternal plasma with fetal fractions greater than 3.7% in more than three fourths of cases after an ultrasonographic gestational age of 8 weeks. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, www.clinicaltrials.gov, NCT01916928. LEVEL OF EVIDENCE: III.

Immunoreactivity of Stat5 phosphorylated on tyrosine as a cell-based measure of Bcr/Abl kinase activity.

BACKGROUND: Stat5(1) (Signal Transducer and Activator of Transcription 5) is normally phosphorylated and activated by Janus kinases. In cells transformed with BCR/ABL, Stat5 is constitutively activated by promiscuous phosphorylation. Cytometry of intracellular antigens can be used to evaluate cell treatments affecting gene expression, because it precisely provides the fraction of affected cells and the quantitative change in expression. Here, we asked whether we could measure a phosphorylated epitope on Stat5 by cytometry, and whether that measurement would respond to Bcr/Abl inhibition. METHODS: Chronic myelogenous leukemia (CML) cell lines or control Bcr/Abl-negative cells were treated or not with imatinib mesylate, fixed and permeabilized with formaldehyde followed by methanol; reacted with rabbit polyclonal and mouse monoclonal antibodies against an epitope including tyrosine 694 of Stat5a (pSTAT5); reacted with antibodies that mark mitotic cells; counterstained with secondary fluorescent antibodies and 4',6-diamidino-2-phenylindole (DAPI); and then subjected to flow cytometry. Western blotting was performed with pSTAT5 and Stat5 antibodies. RESULTS: Optimal fixation and staining parameters were established for pSTAT5 antibodies with K562 cells. These cells displayed high levels of immunoreactivity with pSTAT5 probes that could be inhibited uniformly with imatinib mesylate in a dose-response and time-dependent manner. The IC50 for downregulation of pSTAT5 immunoreactivity for K562 cells by cytometry was approximately 70 nM. The inhibition half-time was approximately 1 min. At micromolar doses this reactivity remained minimal for up to 7 days. Cultured cells also displayed a population of negative cells that increased under conditions related to cessation of cell growth (media nutrient depletion). This study also showed quantitatively that a rabbit polyclonal antibody that cross-reacted with an additional epitope could be used successfully as a measure of Bcr/Abl activity. CONCLUSION: We have developed a sensitive cytometric assay for Bcr/Abl kinase activity in human hematopoietic cell lines.