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Biomedical subjects

C L Wagner

Publications and source records attributed to C L Wagner.

At least 19 recordsLinked to original sources
PubMed2006

Ultrasonographic and radiographic results from a two-year controlled trial of immediate or one-year-delayed addition of infliximab to ongoing methotrexate therapy in patients with erosive early rheumatoid arthritis.

OBJECTIVE: To compare the impact of immediate and delayed introduction of anti-tumor necrosis factor therapy on inflammation and structural damage in methotrexate (MTX)-treated patients with early rheumatoid arthritis (RA). METHODS: Twenty-four patients with erosive early RA (duration < 3 years) who were receiving MTX were randomized to receive infliximab 5 mg/kg or placebo infusions at weeks 0, 2, and 6, and then every 8 weeks through week 46. Beginning at week 54 and thereafter, all patients received infliximab 5 mg/kg. Metacarpophalangeal joints were scanned using high-frequency ultrasonography and power Doppler imaging. Radiographs were evaluated using the modified Sharp/van der Heijde scoring system. RESULTS: From baseline to week 54, total synovial thickness was significantly improved in the infliximab + MTX group compared with the placebo + MTX group (median reduction 95.8% versus 37.5%; P = 0.005), as was the total color Doppler area (CDA; vascularity assessment) (median reduction 100% and 47.1%, respectively; P = 0.025). From week 0 to week 110, no significant between-group difference was observed in the change from baseline for total synovial thickening or the total CDA. At week 54, greater progression in the Sharp/van der Heijde score was apparent in patients receiving placebo + MTX compared with those receiving infliximab + MTX. Although radiographic progression in the placebo + MTX group was greatly reduced in the second year (after initiation of infliximab therapy), marked differences were observed between the infliximab + MTX group (median change in the Sharp/van der Heijde score 4.0) and the placebo + MTX group (median change 14.5) from baseline to week 110 (P = 0.076). CONCLUSION: The results indicate that the efficacy of 2 years of combination therapy with infliximab + MTX for inhibiting cumulative structural damage was superior to that of 1 year of treatment with MTX alone followed by the addition of infliximab.

Antibodies, Monoclonal↗
PubMed2004

Comparison of ultrasonographic assessment of synovitis and joint vascularity with radiographic evaluation in a randomized, placebo-controlled study of infliximab therapy in early rheumatoid arthritis.

OBJECTIVE: To investigate sensitive ultrasonographic imaging methods for detection of synovial thickness and vascularity to discriminate between patients with early rheumatoid arthritis (RA) receiving infliximab + methotrexate (MTX) versus placebo + MTX over 18 weeks, and to compare the relationship between synovial thickening and vascularity at baseline and radiologic damage to joints of the hands and feet at 54 weeks. METHODS: Patients with early RA (duration <3 years) receiving stable dosages of MTX were randomly assigned to receive blinded infusions of 5 mg/kg infliximab (n = 12) or placebo (n = 12) at weeks 0, 2, 6, and then every 8 weeks until week 46. At baseline and week 18, clinical assessments were performed, and metacarpophalangeal joints were assessed by high-frequency ultrasonography and power Doppler ultrasonography measurements. Radiographs of the hands and feet taken at baseline and at 54 weeks were evaluated using the van der Heijde modification of the Sharp method (vdH-Sharp score). RESULTS: Using changes in the total vdH-Sharp score over 54 weeks and changes in synovial thickening and joint vascularity at 18 weeks, we were able to distinguish those patients receiving infusions of infliximab + MTX from those receiving placebo + MTX. Sonographic measurements of synovial thickening and vascularity at baseline in the placebo + MTX group demonstrated clear relationships with the magnitude of radiologic joint damage at week 54. Infliximab + MTX treatment abolished these relationships. CONCLUSION: The delay or reversal of inflammatory and joint-destructive mechanisms in patients with early RA was already apparent following 18 weeks of treatment with infliximab + MTX and was reflected in radiologic changes at 54 weeks.

Antibodies, Monoclonal↗
PubMed2003

Consequences of immunogenicity to the therapeutic monoclonal antibodies ReoPro and Remicade.

The clinical consequences of immune antibodies generated to abciximab (ReoPro) and infliximab (Remicade) are described. Abciximab, a chimaeric Fab fragment that binds to the beta3 integrin of the GPIIb/IIIa and alphavbeta3 receptors on human platelets, is approved in the US and Europe for use in percutaneous coronary intervention (PCI) to prevent cardiac ischaemic complications. The effects of induced antibodies upon the safety and efficacy of repeat administration of abciximab have been evaluated in the ReoPro Re-administration Registry Study, in which 5.7% of patients were HACA positive before re-treatment. An interim evaluation of 1000 patients has indicated that re-administration of abciximab can be accomplished in the setting of PCI with an acceptable safety and efficacy profile. Infliximab is a chimaeric IgG1 antibody specific for human TNFalpha, and is approved in the US and Europe for the acute treatment of the signs and symptoms of Crohn's disease and for the chronic treatment of rheumatoid arthritis (RA). The incidence of antibodies to infliximab is reported to be approximately 10%; however, an inverse dose-immunogenicity relationship was observed, indicating that higher doses of infliximab (> or = 3 to 10 mg/kg) could reduce the incidence of immune antibodies. The induction of immune antibodies could also be reduced by concomitant administration of low-dose methotrexate and other immunosuppressant agents. Although antibodies to infliximab appeared to be associated with lower serum infliximab concentrations and a slightly higher incidence of infusion reactions, these immune antibodies were generally not associated with a reduction in clinical efficacy. In addition, the antibodies induced to infliximab are specific for infliximab, and do not cross-react with other currently available therapeutic antibodies.

Abciximab↗
PubMed2001

Anti-tumor necrosis factor-alpha antibody limits heart failure in a transgenic model.

BACKGROUND: - Tumor necrosis factor (TNF)-alpha has been implicated in the pathophysiology of congestive heart failure. A strain of transgenic mice (TNF1.6) with cardiac-specific overexpression of TNF-alpha develop congestive heart failure. METHODS AND RESULTS: To determine the effect of anti-TNF-alpha therapy in this model, we studied 3 groups: TNF1.6 mice treated with saline, wild-type mice treated with saline, and TNF1.6 mice treated with TNF-alpha neutralizing antibody (cV1q) from 6 to 12 weeks of age. We used echocardiography to compare cardiac hypertrophy, function, and catecholamine response at 12 weeks of age versus baseline (6 weeks). cV1q treatment did not limit cardiac hypertrophy, but it significantly improved basal fractional shortening and responsiveness to beta-adrenergic stimulation, and it limited development of cardiac dilation. CONCLUSIONS: Blockade of TNF-alpha bioactivity by antibody therapy may both preserve cardiac function and partially reverse pathological changes in congestive heart failure.

Adrenergic beta-Agonists↗
PubMed2001

The effect of pasteurization on transforming growth factor alpha and transforming growth factor beta 2 concentrations in human milk.

Transforming growth factor alpha (TGF-alpha) and beta 2 (TGF-beta2) are present in human milk and are involved in growth differentiation and repair of neonatal intestinal epithelia. Heat treatment at 56 degrees C has been shown effective for providing safe banked donor milk, with good retention of other biologically active factors. The purpose of our study was to determine the effect of heat sterilization on TGF-alpha and TGF-beta2 concentrations in human milk. Twenty milk samples were collected from 20 lactating mothers in polypropylene containers and frozen at -20 degrees C for transport or storage. Before heat treatment by holder pasteurization, the frozen milk was thawed and divided into 1-mL aliquots. All samples were heated in an accurately regulated water bath until a holding temperature was achieved, then held for 30 minutes using constant agitation. Holding temperature ranged from 56.5 degrees C to 56.9 degrees C. The milk was then stored at 4 degrees C overnight for analysis the following day. The concentration of TGF-alpha was measured by radioimmunoassay. Mean concentration +/- SD of TGF-alpha in raw milk samples was 119+/-50 pg/mL, range 57 to 234. The mean concentration +/- SD of TGF-alpha in heat treated samples was 113+/-50 pg/mL, range 51 to 227. TGF-alpha concentration was minimally affected by pasteurization, with an overall loss of 6.1%. Of 19 samples, 4 had increased and 15 had decreased concentrations after pasteurization (mean percent SEM: 94%+/-7% of raw milk, range 72%+/-107%). The concentration of acid-activated TGF-beta2 was measured by enzyme-linked immunosorbent assay. Mean concentration +/- SD of TGF-beta2 in raw milk samples was 5624+/-5038 pg/mL, range 195 to 15480. The mean concentration +/- SD of TGF-beta2 in heat-treated samples was 5073+/-4646 pg/mL, range 181 to 15140. TGF-beta2 survived with relatively little loss (0.6%): of 18 samples, 11 had increased and 7 had decreased concentrations after pasteurization (mean percent +/- SEM: 99.4+/-6.7% of raw milk, range 79%-120%). In conclusion, both TGF-alpha and TGF-beta2 were well-preserved in whole milk after holder pasteurization at 56.5 degrees C. The relative increase in growth factor concentration in some of the samples may be attributable to the release of that factor from the cellular and/or fat compartments into the aqueous fraction of human milk. These findings have implications regarding use of donor milk as an alternate source of growth factors and cytokines for the newborn gut when mother's milk is unavailable.

Epithelial Cells↗
PubMed2000

Early discharge with home antibiotic therapy in the treatment of neonatal infection.

OBJECTIVE: To determine the feasibility and cost of home antibiotic therapy for a select group of neonates. METHODS: A cohort of neonates at a university hospital who met criteria for home antibiotic therapy at discharge were prospectively followed (November 1995 to October 1997) for type and duration of antibiotic therapy as well as for hospital readmission. RESULTS: During the study period, 95 infants diagnosed with sepsis, presumed sepsis, pneumonia, or uncomplicated meningitis (having received > 10 days of in-hospital therapy) met prior, established, criteria for home antibiotic therapy. The mean +/- SD birth weight of the cohort was 3160 +/- 526 gm, with a mean gestational age of 38.4 +/- 2.1 weeks. A total of 59 infants (62%) received antimicrobial therapy for a clinical presentation consistent with sepsis or presumed sepsis, and 24 infants (25%) were treated for pneumonia. Ampicillin and gentamicin were prescribed for 56% of the cohort, and ceftriaxone was prescribed for 21% of the cohort. Four of those infants were switched from intravascular ampicillin/gentamicin therapy to intramuscular ceftriaxone after discharge due to loss of intravascular access. With a bilirubin level of > 8, four additional infants were changed from ceftriaxone back to ampicillin and gentamicin to complete coverage. The mean age at discharge was 5.2 days, with a mean hospitalization cost of $6121 for that period. There were no rehospitalizations or emergency department visits secondary to a worsening clinical course. CONCLUSION: In this cohort of neonates who met early discharge and defined home antibiotic therapy criteria, there were no serious complications or treatment failures reported; in addition, there were fewer costs compared with continued inpatient treatment.

Ampicillin↗
PubMed2000

Effect of human milk fortifier on the immunodetection and molecular mass profile of transforming growth factor-alpha.

OBJECTIVE: To determine if the addition of human milk fortifier (HMF) affects the distribution, immunoreactivity, or molecular mass profile of transforming growth factor-alpha (TGF-alpha) within the compartments of human milk. METHODS: Fifteen milk samples were obtained. Each sample was divided into two aliquots; a powdered HMF was added to the first aliquot. TGF-alpha concentration was measured via radioimmunoassay in whole milk and its aqueous and fat fractions +/- HMF. TGF-alpha molecular mass profiles of the samples (v/v) were measured via Western blotting. RESULTS: TGF-alpha concentration (mean +/- SD) in fortified whole milk (15.7 +/- 7.1 pg/100 microl) vs. nonfortified whole milk (14.8 +/- 8.0 pg/100 microl) and in the aqueous fraction of fortified (14.0 +/- 2.7 pg/100 microl) vs. nonfortified (14.0 +/- 3.5 pg/100 microl) did not differ statistically. There was, however, a marked decrease in the concentration of TGF-alpha in the fat fraction of fortified (30.6 +/- 2.8 pg/100 microl) vs. nonfortified (98.0 +/- 6.9 pg/100 microl) milk samples. Western blot for TGF-alpha in whole milk and its separated fractions revealed characteristic bands at 6.5, 12-16, 22, 26-30 and 46 kD. HMF alone and HMF with sodium taurocholate had a prominent band at 18 kD and fainter bands at 6.5, 26-30, and 46 kD. While whole and aqueous milk samples with HMF also consistently showed the 18-kD band, in 8/15 fat fraction samples with HMF the 18-kD band was nondetectable and was only faintly detectable in the remaining 7/15 samples. CONCLUSIONS: It appears that HMF differentially alters the biochemical profile of human milk with regard to TGF-alpha concentration and molecular mass profile. What effect this alteration in human milk biochemistry has on neonatal gut function remains unknown.

Animals↗
PubMed1999

Detection of intracellular tumor necrosis factor alpha in stimulated fetal cells.

BACKGROUND: It is unknown if immature fetal cells produce tumor necrosis factor (TNF) alpha in the same manner that adult cells do. The aim of this study was to determine the feasibility of early detection of intracellular TNF produced by circulating human monocytes (Mo) and lymphocytes (Ly) using flow cytometry and to compare the stimulation profiles of mature and fetal cells. MATERIAL AND METHODS: Fetal umbilical cord blood (n = 10) and adult volunteer blood (n = 10) were obtained. In vitro stimulation with endotoxin (LPS) and ionomycin-PMA was performed. Brefeldin A was added to prevent extracellular transport of TNF. Cell type was determined by using CD-14 marker separating monocyte and lymphocyte populations. Anti-human TNF monoclonal antibody was used to detect intracellular TNF by flow cytometry analysis. RESULTS: Thirty to sixty thousand cells were analyzed per sample. Average TNF expression of stimulated fetal Mo was 28.2%, and that of fetal Ly was only 1.1%. Adult stimulated Ly had an average TNF expression of 31.9%, and adult Mo, 29.6% (P < 0.05 for adult Ly vs fetal Ly). CONCLUSION: TNF flow cytometry analysis allows assessment of individual cell types and their ability to produce that cytokine. Fetal cells are able to produce TNF when stimulated, but the stimulation profile of Ly differs from that of adult samples. This observation may be of clinical importance in evaluating the response of immature cells to a septic stimulus. Flow cytometry is reliable, reproducible, quick, and easily obtained from a small sample of peripheral blood. Clinical use will be applicable once appropriate controls are developed, as reported in this study.

Adult↗
PubMed1999

The use of hypothermia: a role in the treatment of neonatal asphyxia?

Perinatal asphyxia remains one of the most devastating neurologic processes. Although the understanding of the pathophysiology after perinatal asphyxia is extensive, there are few therapeutic interventions available to prevent or even mitigate the devastating process that unfolds after injury. The search for a safe and efficacious therapy has prompted scientists and clinicians to consider various promising therapies. One such therapy is therapeutic hypothermia. On the basis of adult, pediatric, and animal research, there is increasing evidence to suggest that therapeutic hypothermia may be an effective intervention to lessen the secondary neuronal injury that ensues after a hypoxic-ischemic insult. In this article the historic and modern-day uses of therapeutic hypothermia are first reviewed. The pathophysiology of neonatal asphyxia is examined next, with emphasis on the changes that occur when therapeutic hypothermia is implemented. Potential side-effects of the therapy in the neonate and the debate over systemic vs selective hypothermia are discussed. Lastly, although hypothermia as a potential treatment modality for neonates with hypoxic-ischemic encephalopathy is supported by numerous studies, the need for well-designed multicenter trials with detailed patient entry criteria and therapeutic conditions is emphasized.

Adult↗
PubMed1999

Rates of successful vaginal delivery after cesarean for patients with private versus public insurance.

OBJECTIVE: To examine the effect of insurance status on method of delivery while controlling for clinical and sociodemographic characteristics of women delivering at a single medical center. STUDY DESIGN: Sociodemographic and clinical characteristics of 878 women, who delivered their infants between 1985 and 1991 at a university hospital, were analyzed. Women were included if their previous infant was delivered by cesarean section and if the current pregnancy was > or = 37 weeks' gestation at nonemergent delivery with insurance status clearly specified. The outcome of interest was the rate of successful vaginal births after cesarean (VBAC) delivery by insurance status. RESULTS: The trial of labor rate for the cohort, defined as the rate of VBAC or cesarean deliveries following labor, was 55%; 61% were vaginal deliveries. Significant differences with regard to insurance status and several maternal factors were noted between trial of labor and cesarean section--no labor groups. After controlling for potentially confounding variables, the Medicaid/indigent group was more likely than the privately insured group to undergo a trial of labor (odds ratio, 1.5; 95% confidence interval, 1.1 to 2.4). Of women who underwent a trial of labor, after controlling for other characteristics, the Medicaid/indigent group was more likely than the privately insured group to deliver vaginally (odds ratio, 1.9; 95% confidence interval, 1.1 to 3.2). CONCLUSION: After controlling for other covariables, women with a history of a prior cesarean section with Medicaid/indigent insurance were more likely than privately insured women to attempt a trial of labor, and subsequently, to deliver vaginally, given that a trial of labor was attempted.

Adult↗
PubMed1999

The breast or the bottle? Determinants of infant feeding behaviors.

Although various trends have placed breastfeeding in and out of vogue, in the twentieth century the greater availability of human milk substitutes mandates that a woman choose her infant's feeding method. It appears that intrapsychic factors or life experiences, as well as certain social conditions, influence that choice. For example, the economic state of society historically has had significant impact on the role of women and the value placed on woman's unique biologic contributions. Likewise, personality and attitudinal factors also may act as potential mediators of observed differences between lactating and nonlactating mothers in their mother-infant interactions. Finally, once the decision to breastfeed or bottle-feed has been made and carried through, additional physiologic mechanisms may mediate conscious behavioral intentions. The phenomenon of human lactation, then, is sensitive to a variety of interrelated factors that can be grouped as follows: (1) individual personality, (2) social forces, and (3) psychophysiologic mechanisms. An in-depth understanding of the specific factors that affect a woman's decision to breastfeed will have far-reaching implications for future educational and interventional programs.

Attitude↗
PubMed1998

Pharmacodynamic profile of short-term abciximab treatment demonstrates prolonged platelet inhibition with gradual recovery from GP IIb/IIIa receptor blockade.

BACKGROUND: The glycoprotein (GP) IIb/IIIa receptor antagonist abciximab is approved for use in high-risk percutaneous coronary interventions. The purpose of the present study was to establish the pharmacodynamic profile and platelet-bound life span of abciximab. METHODS AND RESULTS: The pharmacodynamics of abciximab (inhibition of ex vivo platelet aggregation and GP IIb/IIIa receptor blockade) were measured in 41 individuals who were randomized to receive a 0.25-mg/kg bolus and a 12-hour infusion of either 10 microg/min (EPIC regimen) or 0.125 microg x kg(-1) x min(-1) (EPILOG regimen) of the antiplatelet agent. At extended times, the amount and distribution of platelet-bound abciximab were monitored by flow cytometry. The EPIC and EPILOG infusion regimens exhibited equivalent blockade of both GP IIb/IIIa receptors and platelet aggregation throughout the duration of abciximab treatment. Flow cytometry revealed a single, highly fluorescent platelet population during treatment, consistent with complete saturation and homogeneous distribution of abciximab on circulating platelets. For 15 days after treatment, the fluorescence histograms remained unimodal with gradually diminishing fluorescence intensity, indicating decreasing levels of platelet-bound abciximab. At 8 and 15 days, which exceeds the normal circulating life span of platelets, median relative fluorescence intensity corresponded to 29100 (29% GP IIb/IIIa receptor blockade) and 13300 (13% GP IIb/IIIa receptor blockade) abciximab molecules bound per platelet, respectively. CONCLUSIONS: These results are consistent with continuous reequilibration of abciximab among circulating platelets and may explain the gradual recovery of platelet function and long-term prevention of ischemic complications by abciximab after coronary intervention.

Abciximab↗
PubMed1998

The impact of prenatal drug exposure on the neonate.

Several social or recreational drugs singly or together have demonstrated effects on the fetus and neonate, with those effects extending into adulthood. The use of recreational drugs during pregnancy remains a major health problem, with adverse effects including higher rates of fetal distress and demise, lower Apgar scores, growth retardation, and adverse neurodevelopmental outcome. Ethanol has the most profound effects, with physical stigmata of the drug seen in one third of exposed infants. In children without the affected physical appearance, profound neurodevelopmental sequelae have been demonstrated. Other drugs, such as cocaine, heroin, amphetamines, and nicotine, have been associated with impaired fetal growth and acute withdrawal during the neonatal period. Subsequently, these infants and children have an increased risk for altered neurodevelopment and long-term health status. Long-term follow-up and assessment are essential. The risk of neonatal withdrawal or abstinence syndrome is greatest with narcotic drugs but has been found to occur in neonates following exposure to cocaine, nicotine, and amphetamines. Early treatment with tincture of opium, paregoric, or phenobarbital is crucial. Assessment of the overall health status of the infant should include growth parameters, signs and symptoms of infection (especially hepatitis, syphilis, and immunodeficiency viruses), and neurobehavioral function. Such assessments should not be limited to the newborn period, as neurodevelopmental sequelae may not be manifest until later in infancy and childhood. In addition, evaluation of the social milieu is warranted because of the increased risk for neglect and abuse of drug-exposed infants and children. Early intervention, maternal drug rehabilitation treatment, and parenting classes are frequently prescribed, but their efficacy is variable. Further investigations should study the potential benefits of these recommendations.

Developmental Disabilities↗
PubMed1998

The effect of recombinant TGFalpha, human milk, and human milk macrophage media on gut epithelial proliferation is decreased in the presence of a neutralizing TGFalpha antibody.

OBJECTIVE: An in vitro model was devised to compare the relative effects of recombinant transforming growth factor-alpha (TGFalpha), aqueous human milk, and human milk macrophage (HMM) medium on human fetal small intestinal cell (FHs-74) proliferation. METHODS: Recombinant TGFalpha at increasing concentrations (range 0.01-1,000 ng/ml media), the aqueous fraction of human milk (AHM), or HMM medium was added to FHs-74 cells in the presence or absence of a neutralizing TGFalpha antibody (1 microgram/ml medium). At 24 h, cell proliferation was measured and expressed as percent control. The experimental variables were (1) activators of cell growth (TGFalpha, AHM, and HMM medium); (2) increasing concentrations of TGFalpha, and (3) neutralizing antibody to TGFalpha. The dependent variable for all experiments was cell proliferation. RESULTS: Significant effects for growth stimulators and TGFalpha concentration as measured by cell proliferation were found. Specifically, there was a dose-dependent effect of TGFalpha on cell proliferation to the 5-ng/ml concentration, with a plateau reached in cell proliferation at higher concentrations. The stimulatory effect of TGFalpha was decreased in the presence of TGFalpha antibody (mean +/- SD 22 +/- 7. 1% decline, p < 0.001). In the presence of TGFalpha antibody, there was a 25 +/- 3.1% decline in HM-stimulated growth (p < 0.004), and a 27.6 +/- 3.2% decline in HMM medium-stimulated growth (p < 0.001). CONCLUSIONS: Neutralization of recombinant TGFalpha and that present in human milk and HMM medium by TGFalpha antibody led to a consistent decrease in in vitro human fetal small intestine epithelial proliferation without affecting cell viability. These results support the hypothesis that TGFalpha, whether derived from human recombinant sources, human milk or HMM medium has a measurable, trophic effect on in vitro human gut epithelial cells.

Animals↗
PubMed1997

Drug therapy in the nursing mother.

The benefits of breast-feeding on newborn and infant health and for the maternal-infant relationship are well established. It is a failing of health care providers that breast-feeding is too frequently interrupted because of unfounded concerns over the potential adverse effects of concurrent drug therapy in the nursing mother. An overwhelming minority of medications are contraindicated during lactation, while the vast majority are considered either safe to use or the risk versus benefit considerations clearly favor use with continuation of breast-feeding. For those drugs that may pose a potential concern to the newborn, there are practical suggestions and precautions that can be taken to minimize risk. These include such interventions as selection of alternative effective drug regimens with safer profiles in breast-feeding, timing of drug dosing to minimize accumulation in the breast milk, surveillance for newborn or infant symptomatology that may be a sign of toxicity, or even the determination of drug levels in the infant's circulation. With a commitment to the advantages of breast-feeding and the information contained both herein and elsewhere regarding the interaction of medical therapy, pharmacology, and lactation, it is hoped that the health care provider will be empowered both to encourage and counsel more effectively the parturient who wishes to breast-feed but is concerned about ongoing drug therapy during lactation.

Adult↗
PubMed1996

Analysis of GPIIb/IIIa receptor number by quantification of 7E3 binding to human platelets.

A large number of glycoprotein (GP) IIb/IIIa receptors are present on the surface of platelets. Studies to define precisely the number of GPIIb/IIIa receptors using specific monoclonal antibodies (MoAbs) or fibrinogen binding have, however, yielded varying estimates of receptor number. To refine the quantitative estimation of GPIIb/IIIa receptors on resting platelets, we have used the MoAb 7E3, which has high affinity for GPIIb/IIIa. Quantitative binding studies were performed using radiolabeled conjugates of 7E3 IgG, as well as fragments and derivatives of 7E3. For platelets obtained from any single individual, the numbers of 7E3 F(ab')2 and IgG molecules bound per platelet were equivalent (approximately 40,000), whereas the number of Fab molecules bound per platelet was consistently approximately twofold higher (approximately 80,000). To investigate the basis of the quantitative disparity in binding of intact 7E3 and 7E3 F(ab')2 versus 7E3 Fab, we studied the binding of a newly constructed, bispecific (Fab')2 molecule containing only a single 7E3 combining site. Because this construct bound to the same extent as the Fab species, the larger size of the intact 7E3 and 7E3 F(ab')2 molecules could not explain the reduced number of molecules that bound per platelet compared to the Fab fragment. Rather, it appears that the valency of the antibody is the critical factor determining the number of antibody molecules bound per platelet. Thus, we conclude that the binding of 7E3 Fab corresponds most closely with surface GPIIb/IIIa number and that the number of GPIIb/IIIa receptors is approximately 80,000 per platelet.

Antibodies, Bispecific↗