Preimplantation single cell analyses of dystrophin gene deletions using whole genome amplification.

Primer extension preamplification (PEP) increases the scope and capacity of single cell genetic diagnosis by generating sufficient template to perform multiple subsequent DNA analyses using the polymerase chain reaction. We report the simultaneous analysis of single cells at five commonly deleted dystrophin exons and at the ZFX/ZFY loci. Ninety three percent of PEP reactions with single amniocytes, chorionic villus cells and blastomeres were successful, and a blinded analysis of single lymphoblasts from affected males resulted in 93% diagnostic accuracy, demonstrating its applicability in preimplantation prevention of Duchenne muscular dystrophy. Transfer of unaffected male embryos and improved diagnostic reliability are achieved with the ability to perform replicate multilocus analyses from the same blastomere.

Detection of actin-binding proteins in human platelets by 125I-actin overlay of polyacrylamide gels.

Actin-binding proteins have been identified in human platelets with a gel-overlay technique that uses 125I-G-actin. Platelet proteins were separated on SDS polyacrylamide gels using the buffer system of Laemmli (1970, Nature [Lond.] 227:680-685). The proteins were fixed in the gels with methanol-acetic acid, the SDS was washed out, and the proteins were renatured. The gels were incubated with 125I-G-actin from rabbit skeletal muscle that was radiolabeled with 125I according to the method of Bolton and Hunter (1973, Biochem. J. 133:529-538) and has been shown to retain biological activity. After nonspecifically bound radioactivity was washed out, gels were dried and processed for autoradiography. The 125I-G-actin binds to several proteins in human platelets, platelet extracts, and the particulate fraction. Control experiments demonstrate that the 125I-G-actin can be displaced by use of increasing amounts of unlabeled actin, that the binding is stable to 0.6 M NaCl, and that preheating the 125I-G-actin to 90 degrees C for 3 min eliminates all binding. Prominent 125I-G-actin-binding activities were present at Mr 90,000 and 40,000. The binding to the 90,000 Mr protein appears to be at least partially Ca++ sensitive, whereas the binding to the 40,000 Mr protein does not. 125I-G-actin bound to proteins in the SDS gels can be fixed in situ and compared directly with the stained gel. This technique should prove generally useful in identification and purification of some actin-binding proteins from cells and tissues.

A daily neural signal for luteinizing hormone release in the untreated ovariectomized rat: changes in gonadotropin-releasing hormone content of the preoptic area and hypothalamus throughout the day.

Gonadotropin-releasing hormone (GnRH) content of preoptic areas (POA) and hypothalami, and serum gonadotropins of rats ovariectomized six weeks earlier were measured throughout the day in two experiments. In the first, rats were decapitated at 2 h intervals between 0800 and 1800 h. The entire preoptic-hypothalamic region was removed and extracted for radioimmunoassay (RIA) of GnRH. Serum gonadotropins measured by RIA were highly variable but mean concentrations were not significantly different throughout the day. However, preoptic hypothalamic content of GnRH declined markedly between 1000 and 1200 h. In the second experiment, 75 rats were divided into three groups and were untreated or were implanted sc with empty Silastic capsules or capsules containing estradiol-17beta (E2). Two days later, groups of five rats from each of the three treatment groups were decapitated at 0800, 1100, 1400, 1700 and 2000 h. The preoptic area was separated from the hypothalamus by a transverse cut at the caudal aspect of the optic chiasm. POA and hypothalamic content of GnRH correlated well (r=0.74, P less than 0.001, n=75). Two-way analysis of variance failed to reveal any effect of treatment on the GnRH content in either the POA or hypothalamus. GnRH content of both regions decreased significantly between 1100 and 1700h regardless of whether E2 was administered. In striking contrast, gonadotropin surges occurred in the late afternoon only in the E2-treated rats. Serum GnRH was undetectable (less than 5 pg/ml) in all groups of animals. These experiments demonstrate that in the untreated ovariectomized rat GnRH content of the POA and hypothalamus decreases during the early afternoon. This study supports the concent of a daily neural signal for LH release and that E2 is necessary for expression of the daily LH surge in the ovariectomized rat.

Characterization of prostaglandin E2 binding to uterine membranes from baboon, rabbit and tree shrew (Tupaia belangeri).

Membrane preparations (100,000 g pellet) of rabbit, baboon and tree shrew (Tupaia belangeri) uteri were studied for binding of [3H]prostaglandin E2 ([3H]PGE2). Unbound [3H]PGE2 was separated by filtration through Whatman GF/F filters. Non-specific binding was determined by the amount of radioactivity associated with the filters in the presence of a 100-fold excess of radioinert PGE2. PGE2 bound to membranes could be displaced by some other prostaglandin (PG) molecules: PGE1, 16,16-dimethyl-PGE2, PGA1 and PGF2 alpha, but not by 6-keto-PGF1 alpha, PGD2 or arachidonic acid. No PGE2 binding was detected using either membrane ghosts from red blood cells or liposomes. Apparent equilibrium of the binding was reached by 60 min. There was no difference in dissociation constant (Kd) values between rabbits of different reproductive stages (mean range +/- S.E.M. was from 4.6 +/- 0.3 to 5.5 +/- 1.0 nM), but pregnant baboons showed a significantly lower value (3.3 +/- 0.4 nM) than did cyclic animals (12.0 +/- 2.0 nM). Binding capacity (Bmax) values, in contrast, were different only between oestrous rabbits and other reproductive stages. The small amounts of Tupaia tissue only permitted estimates of the Kd and Bmax values to be made; these were 3.8 nM and 499 fmol/mg protein for oestrous animals and 5.4 nM and 674 fmol/mg protein for animals on day 7 of pregnancy, assuming only one class of sites. The present results demonstrate the presence of specific binding sites for PGE2 in uteri from several species.

Ascorbic acid supplement use and the prevalence of gallbladder disease. Heart & Estrogen-Progestin Replacement Study (HERS) Research Group.

To investigate the relation of ascorbic acid supplement use to gallbladder disease and cholecystectomy, we conducted a cross-sectional analysis of baseline from 2744 postmenopausal women, aged 44-79 years, enrolled in the Heart & Estrogen-progestin Replacement Study (HERS), a secondary coronary heart disease prevention trial. A total of 629 HERS participants (23%) reported a history of gallbladder disease. Of these, 508 (19%) also reported a history of cholecystectomy. In bivariate models, ascorbic acid supplement use was associated with a decreased prevalence of gallbladder disease [odds ratio (OR)=0.74; 95% confidence interval (CI), 0.57, 0.96] and a trend toward a decreased prevalence of cholecystectomy (OR=0.77; 95% CI, 0.58, 1.02). Because we detected significant interactions between ascorbic acid supplement use and alcohol consumption, multivariate analyses were performed stratified by drinking status. After adjustment for potential confounding variables, use of ascorbic acid supplements among drinkers was associated with a decreased prevalence of gallbladder disease (adjusted OR=0.50; 95% CI, 0.31, 0.81) and cholecystectomy (adjusted OR=0.38; 95% CI, 0.21, 0.67). Use of ascorbic acid supplements among non-drinkers was not significantly associated with either prevalence of gallbladder disease or cholecystectomy. Further study is necessary to confirm our findings and, specifically, to examine the combined effects of ascorbic acid and alcohol on cholesterol metabolism.

Laparoscopic tubal sterilization under local anesthesia in women with cyanotic heart disease.

Laparoscopic tubal sterilization under local anesthesia with intravenous sedation has been shown to be a safe procedure. However, the use of laparoscopy in patients with cyanotic cardiovascular disease is controversial and is generally contraindicated. Five women were referred with uncorrectable cyanotic heart disease and pulmonary hypertension. The mean preoperative arterial oxygen pressure was 56.2 +/- 5 mmHg (N = 5). After cardiology and cardiovascular anesthesia consultation and clearance, the patients underwent laparoscopic sterilization with Silastic rings under local anesthesia using direct trocar entry. Continuous hemodynamic monitoring and pulse oximetry were employed. The patients were kept in the intensive care unit or the hospital for 24 hours for monitoring, and all did well. This hospital for 24 hours for monitoring, and all did well. This small retrospective series demonstrates that laparoscopic sterilization under local anesthesia is a sterilization technique that may be suitable and safe for such patients when appropriate monitoring is performed. Tubal sterilization may be the contraceptive method of choice in women with heart disease when pregnancy is contraindicated.

GnRH agonist exposure during the first trimester of pregnancy.

OBJECTIVE: To evaluate pregnancy outcome in infertility patients with unrecognized exposure to a GnRH agonist in the first trimester. METHODS: Five women were given GnRH agonist before controlled ovarian hyperstimulation for in vitro fertilization cycles. The medication was administered in a dose of 0.5 mg/day, with drug exposure beginning on cycle day 21. The duration of exposure in all patients was 14-21 days. Thus, all five women received the medication at 3-6 weeks' estimated gestational age by menstrual dating. Pregnancy tests were not performed before the first injection of the GnRH agonist. RESULTS: Three of the five pregnancies progressed to term without complication, and normal healthy infants were delivered. Missed abortion occurred in one pregnancy, and another ended in induced abortion at 13 weeks because of trisomy 18. CONCLUSIONS: This experience suggests that despite manipulation of the hypothalamic-pituitary-ovarian axis by administration of GnRH agonist in the first trimester of pregnancy, normal pregnancies can result. Pregnancies in these patients should not be terminated because of drug exposure alone.

Physiologic estradiol replacement therapy and cardiac structure and function in normal postmenopausal women: a randomized, double-blind, placebo-controlled, crossover trial.

OBJECTIVE: To assess the effect of estradiol (E2) replacement therapy on cardiac structure and function in healthy postmenopausal women. METHODS: We conducted a randomized, double-blind, placebo-controlled, crossover study of 31 healthy postmenopausal female volunteer study subjects (55-65 years) using 12 weeks of micronized E2 replacement therapy (2 mg/day). Echocardiography and Doppler techniques were used to assess the cardiac effects of E2 at rest and during graded bicycle ergometry. RESULTS: Crossover analysis demonstrated no carryover effects of estrogen treatment (which increased serum E2 15-fold to 37.6 pmol/L) on the cardiac characteristics measured. Estradiol treatment did not affect measurements of systolic function, diastolic function, left ventricular mass, or pulmonary artery pressure at rest or during bicycle ergometry. Left ventricular end-diastolic volume at rest was slightly higher with E2 treatment (P = .03). However, this change was not reflected by changes in stroke volume, ejection fraction, or cardiac output. CONCLUSIONS: Estrogen replacement therapy, which results in physiologic serum concentrations, does not affect cardiac structure or function in normal postmenopausal women after 12 weeks of treatment.

Effect of postmenopausal estrogen replacement on circulating androgens.

OBJECTIVE: To determine the effect of estrogen replacement therapy (ERT) on serum androgen levels in postmenopausal women. METHODS: We measured serum dehydroepiandrosterone (DHEA), DHEA-sulfate, testosterone, estradiol (E2), LH, FSH, and sex hormone binding globulin in 8:00 AM fasting serum samples from a previous randomized, blinded, placebo-controlled crossover study in which 28 postmenopausal women (27 naturally menopausal) were given 2 mg/day of oral micronized estradiol. The treatment arms were 12 weeks with a 6-week washout. RESULTS: Estrogen replacement therapy raised mean (+/- standard error of the mean [SEM]) serum E2 from 8.7 +/- 1.0 to 117 +/- 18.7 pg/mL (P < .001 from baseline). Concurrently, mean (+/- SEM) DHEA-sulfate fell from 67.3 +/- 9.6 to 52.1 +/- 6.4 micrograms/dL (P < .001), and mean (+/- SEM) testosterone fell from 16.1 +/- 2.4 to 9.4 +/- 1.4 ng/dL (P = .006). Both FSH and LH declined significantly. Sex hormone binding globulin increased by 160% with ERT (P < .001). CONCLUSION: Menopausal ERT decreases serum androgen levels, decreasing DHEA-sulfate and testosterone by 23% and 42%, respectively. Whereas the decline in testosterone is likely due to decreased LH-driven ovarian stromal steroidogenesis, the declining levels of DHEA-sulfate also may imply a direct adrenal effect of estrogen. Bioavailable testosterone likely is reduced even more profoundly because sex hormone binding globulin is increased 160% by estrogen. Thus, menopausal ERT may induce relative ovarian and adrenal androgen deficiency, creating a rationale for concurrent physiologic androgen replacement.

The effects of a long-acting progestin on the hypothalamic-pituitary-ovarian axis in women with normal menstrual cycles.

This study was performed to determine how a long-acting, slow-release preparation of norethindrone (NET) affects the hypothalamic-pituitary-ovarian axis of normal ovulatory women. Ten women were studied during the luteal phase of their menstrual cycle, and again at six and twelve weeks following intramuscular administration of 100 mg NET microencapsulated in poly-D,L-lactide-co-glycolide. Serial LH samples, serum E, P, and NET were followed by a GnRH stimulation test. Compared to luteal phase values, six and twelve weeks of treatment with NET inhibited serum E2 and P while mean serum LH remained unchanged and mean serum FSH increased significantly (p < 0.05). LH pulse frequency after NET treatment was twice the rate (p < 0.01) as that of the luteal phase, whereas LH pulse amplitude was decreased significantly (p < 0.05). Finally, although there was no significant change in pituitary LH secretion in response to GnRH, NET treatment augmented FSH responsiveness to GnRH at the times studied. Preserved pituitary responsiveness to GnRH in NET-treated patients suggests that inhibited ovarian function results in an increase in GnRH pulse frequency but not GnRH pulse amplitude. Since the progestational milieu is maintained in these patients by NET treatment, the decrease in serum E2 may be responsible for the increase in GnRH pulse frequency. The presence of a critical level of E2 may be necessary for progestins to affect the hypothalamic GnRH pulse generator.

A severe puerperal group A streptococcal infection causing a toxic shock-like syndrome.

Toxic shock syndrome (TSS) is classically associated with vaginal recovery of Staphylococcus aureus during menses. In this case a patient presented with fever, rash, abdominal pain and signs of shock, 6 days postpartum. Blood cultures were negative but endometrial cultures were positive for Group A beta-hemolytic streptococcus. This case presents a toxic shock-like syndrome due to streptococcus, (toxic streptococcus syndrome) and points out the importance of culturing these patients for organisms other than Staphylococcus aureus.

Hysterosalpingographic diagnosis of Crohn's disease. A case report.

A woman presented with abdominal pain, weight loss and a pelvic mass. At the time of laparotomy she had a lower abdominal abscess from perforation of the ileum. Two years later she returned with a tender uterus and purulent cervical discharge. A hysterosalpingogram demonstrated an uteroileal fistula secondary to Crohn's disease, and the patient underwent a total abdominal hysterectomy and bilateral salpingo-oophorectomy.

Clinical use of nafarelin in the treatment of leiomyomas. A review of the literature.

OBJECTIVE: To review the efficacy and safety of nafarelin in the treatment of leiomyomas. STUDY DESIGN: A literature review of published clinical trials was conducted. Six studies, including a total of 602 patients with leiomyomas, were reviewed. Patients received intranasal nafarelin, 50-400 micrograms twice daily for three to six months. Vaginal bleeding patterns, leiomyoma and uterine size, surgical conditions and adverse effects were assessed. RESULTS: Nafarelin consistently suppressed estrogen production, reduced leiomyoma and uterine size, and controlled menorrhagia. The significant reduction in uterine bleeding and amenorrhea resulting from administration of nafarelin was associated with a rise in mean hemoglobin concentrations. In addition, nafarelin improved hematologic parameters in women with and without anemia. Nafarelin was well tolerated, although hot flushes were the most commonly reported adverse events. Measured bone mineral density decreased significantly during treatment, although by six to nine months post-treatment, it increased to values not significantly different from baseline. The adverse effects of nafarelin were generally reversible after treatment withdrawal. CONCLUSION: Nafarelin treatment of women with symptomatic leiomyomas effectively decreases uterine bleeding; improves hematologic parameters; manages symptoms of menometrorrhagia, dysmenorrhea and pelvic discomfort; reduces uterine and myoma size; and is well tolerated. Reduction in bone mineral density occurs, but levels return to, or near, baseline levels within six months after treatment.

Prevalence of HIV seropositivity among inner-city adolescents in 1988 and 1992.

The purpose of this study was to document the change in prevalence of human immunodeficiency virus (HIV) infection among sexually active minority teens at a primary and reproductive teen health clinic. Both new and current patients were sampled in a seroprevalence study in 1988 and again in 1992. None of the 1200 adolescents sampled were seropositive for HIV in 1988. Nine of the 1085 adolescents sampled were seropositive in 1992. Five of these 9 teens reported heterosexual contact only as a potential risk factor. Six of these 9 teens were not in school. These results suggest that HIV infection is increasing among Texas urban teens. Because current education programs do not appear to be preventing viral spread, new, focused intervention must be initiated for teenagers. In particular, an effective acquired immunodeficiency syndrome prevention program for out-of-school youth is needed urgently.

Site of action of indomethacin on implantation in the rabbit.

Blastocysts recovered from oil- or indomethacin-treated donor rabbits between 5 1/2 and 6 days after insemination and hCG injection were transferred to oil- or indomethacin-treated recipients between 135 and 147 h after hCG injection. Indomethacin treatment of donor rabbits (10 mg/kg s.c.) given every 6 h during the day before transfer had no effect on subsequent implantation of the blastocysts. However, indomethacin treatment of the recipients (10 mg/kg s.c. every 6 h from 120 to 168 h after hCG) prevented implantation of all transferred blastocysts, although 6 of the 8 rabbits died between Days 9 and 16 of (pseudo)pregnancy. Restriction of the indomethacin treatment of the recipients to only 3 injections of 10 mg/kg s.c. between 128 and 140 h after hCG injection had no effect on the implantation of the transferred blastocysts. It is concluded that indomethacin exerts its inhibitory influence on implantation via an action on the endometrium rather than on the blastocyst.

Acute inhibitory effects of antiserum to gonadotropin-releasing hormone in ovariectomized rats. Evidence for pulsatile secretion of gonadotropin-releasing hormone.

To evaluate the dependence of pulsatile secretion of luteinizing hormone (LH) on gonadotropin-releasing hormone (GnRH), the acute effects of immunoneutralization of endogenous GnRH on plasma LH were compared in ovariectomized rats with phenobarbital treatment and hypophysectomy. Anti-GnRH produced a rapid dose-dependent decrease in plasma LH and LH pulse amplitude. Pulsatile secretion of LH was eliminated in 6 of 12 rats treated with the highest dose of anti-GnRH, but plasma LH was still 50% of control values 3 h after treatment. Frequency of LH pulses was unchanged in animals which had persistence of pulsatile secretion of LH. Phenobarbital eliminated pulsatile secretion of LH transiently. Hypophysectomized rats displayed a striking decrease in plasma LH which could be resolved into two exponential components with half-lives of 16 and 70 min. The initial half-life of plasma LH from untreated rats determined after LH pulses was also 16 min. These studies support the hypothesis that pulses of GnRH induce the pulsatile pattern of plasma LH and may be responsible for all LH secretion in ovariectomized rats.

Cryopreserved mouse embryos can successfully survive biopsy and refreezing.

OBJECTIVE: We examined whether cryopreserved mouse eight-cell embryos could be thawed, biopsied, refrozen, thawed, and grown in vitro and in vivo in two sets of experiments. METHOD: In the in vitro studies, the blastulation rate of cryopreserved embryos which had been thawed-->biopsied-->refrozen-->thawed-->grown 48 hr in vitro were compared with that of sham-operated (zona dissected) and untreated control embryos (twice frozen only). For the in vivo studies, five control embryos were transferred to one horn and five experimental embryos were transferred to the contralateral horn of day 3 pseudopregnant recipient female mice. Recipient mice were sacrificed on day 18. RESULTS: The day 2 blastulation rate was the same for the control, sham-operated, and biopsied embryos when examined in vitro. In the first set of in vivo studies, 42.7% of control and 39.7% of sham-operated embryos that had been transferred implanted, and most embryos progressed to day 18. In the second set, 45.6% (57/125) and 39.7% (49/125), respectively, of the transferred embryos progressed to day 18 fetuses. There were no significant differences in the rate of fetal development in the different groups. CONCLUSIONS: These studies demonstrate that cryopreserved mouse eight-cell embryos can successfully undergo thawing, biopsy, and refreezing. The results suggest that under certain conditions, it may be possible to utilize cryopreservation in strategies involving human genetic diagnosis in the preimplantation period.

Identification of G actin-binding proteins in rat tissues using a gel overlay technique.

Actin-binding proteins were assayed in various tissues using an 125I-actin overlay procedure. Four major G actin-binding proteins of 90000, 65000, 58000 and 40000 Mr have been identified. The 90K protein is present in all tissues and binds labelled actin in a calcium-sensitive manner with binding increasing 3-4-fold in the presence of Ca2+. The distribution of the 58K and 65K protein which are not Ca2+-sensitive was more variable. These proteins were present in different ratios in different tissues. 125I-actin binding to all four actin-binding proteins is specific and can be displaced by preincubation of the gels with unlabelled actin. The interaction of actin with these proteins does not appear to involve ionic forces, since binding is not diminished by varying the salt concentration. Skeletal muscle glycolytic enzymes, the lens crystallins and the histones also bind 125I-actin. This binding cannot be displaced by preincubation with unlabelled actin and is presumably non-specific. The calcium sensitivity of two highly purified actin-binding proteins, the 90K human platelet protein and villin was compared using 125I-actin. The platelet 90K protein binds actin at less than 10(-7) M free calcium, but detectable binding to villin does not occur below 10(-6) M free calcium. The ubiquity of these actin-binding proteins is clear and we conclude that the calcium-sensitive 90K actin-binding protein in all of these tissues is the same as the platelet protein.

A randomized, double-blind, placebo-controlled comparison of the impact of low-dose and triphasic oral contraceptives on follicular development.

OBJECTIVE: This investigation tests the hypothesis that triphasic oral contraceptives are associated with the development of large, persistent ovarian cysts. STUDY DESIGN: Weekly vaginal ultrasonography was used in a randomized, double-blind, placebo-controlled, parallel-group, single-center study that compared the incidence, risk, size, and time to resolution of ovarian follicles in healthy women who took Estrostep or Loestrin oral contraceptives (manufactured by Parke-Davis) or a placebo during three consecutive menstrual cycles. RESULTS: Sixty-three percent of placebo-treated subjects developed follicles greater than 18 mm, compared with 39% and 23% in the Estrostep and Loestrin groups. The risks for each group of developing a large follicle during a single cycle were not different. No dominant follicle persisted for greater than 2 weeks for any subject. CONCLUSION: These results demonstrate that follicular development continues during treatment with oral contraceptives. In addition, the findings fail to support the hypothesis that triphasic oral contraceptives result in persistent ovarian cysts.

PIP: Health practitioners randomly enrolled 48 18-35 year old healthy women into either the group receiving a low dose oral contraceptive (Loestrin, 30 mcg of ethinyl estradiol and 1.5 mg of norethindrone acetate) or the group receiving a triphasic oral contraceptive (Estrostep, 20, 30, and 35 mcg of ethinyl estradiol and 1.5 mg of norethindrone acetate) and followed them for a 4-week control period. They were able to follow only 42-45 women during active treatment. This double blind, placebo controlled study took place at the Baylor College of Medicine in Houston, Texas. Its purpose was to determine whether Estrostep was causally related to development of ovarian cysts. 63% of the women in the placebo group had a follicle 18 mm sometime during treatment compared with 43% of those treated with Estrostep and 25% of those treated with Loestrin. No group was more prone to developing follicular cysts than the other 2 groups. Mean size of these follicles were 21, 21.5, and 23.8 mm for the control, Loestrin, and Estrostep groups, respectively, and were not significantly different. The mean largest follicle sizes were also not significantly different. None of the women experienced a follicle 18 mm for 2 weeks. The results indicated that follicular development persists during oral contraceptive treatment. They also do not support the hypothesis that triphasic oral contraceptives cause continual ovarian cysts.

A DNase I binding/immunoprecipitation assay for actin.

An actin assay which employs the competition between labeled and unlabeled rabbit skeletal muscle actin for DNase I has been developed. Iodination of actin by the method of Bolton and Hunter results in the incorporation of approximately 0.5 mol of 125-iodine/incorporation of approximately 0.5 mol of 125-iodine/mol of actin. This 125I-actin retained the ability to bind to DNase I and inhibit enzymatic activity. The 125I-actin-DNase complex can be precipitated by the addition of a monospecific rabbit antibody to DNase I. The efficiency of this immunoprecipitation step is improved by the use of a second sheep anti-rabbit gamma-globulin. Using this immunoprecipitation assay, there is a linear displacement of the DNase I-bound 125I-actin by rabbit skeletal muscle actin standards or by the actin present in tissue and cell extracts. Using 17.5 ng of DNase I and approximately 500 pg of 125I-actin, 50% inhibition of binding was obtained with 23 ng of unlabeled actin. Reducing the amount of DNase I to 2 ng results in 50% inhibition of binding with 4 ng of unlabeled actin and an increase in the estimated sensitivity of the assay from 1.7 to 0.24 ng. The slopes of the displacement curves generated with both vertebrate and invertebrate non-muscle actins are parallel to rabbit skeletal muscle actin. This observation indicates approximately equal actin-DNase I binding affinities and suggests a high degree of conservation of the actin-DNase I binding site. The assay is useful for measuring the pools of F- and G-actin in a wide range of cells.