Human monoclonal antibody MBL-HCV1 delays HCV viral rebound following liver transplantation: a randomized controlled study.

Rapid allograft infection complicates liver transplantation (LT) in patients with hepatitis C virus (HCV). Pegylated interferon-α and ribavirin therapy after LT has significant toxicity and limited efficacy. The effect of a human monoclonal antibody targeting the HCV E2 glycoprotein (MBL-HCV1) on viral clearance was examined in a randomized, double-blind, placebo-controlled pilot study in patients infected with HCV genotype 1a undergoing LT. Subjects received 11 infusions of 50 mg/kg MBL-HCV1 (n=6) or placebo (n=5) intravenously with three infusions on day of transplant, a single infusion on days 1 through 7 and one infusion on day 14 after LT. MBL-HCV1 was well-tolerated and reduced viral load for a period ranging from 7 to 28 days. Median change in viral load (log10 IU/mL) from baseline was significantly greater (p=0.02) for the antibody-treated group (range -3.07 to -3.34) compared to placebo group (range -0.331 to -1.01) on days 3 through 6 posttransplant. MBL-HCV1 treatment significantly delayed median time to viral rebound compared to placebo treatment (18.7 days vs. 2.4 days, p<0.001). As with other HCV monotherapies, antibody-treated subjects had resistance-associated variants at the time of viral rebound. A combination study of MBL-HCV1 with a direct-acting antiviral is underway.

Correlation of the Km(1) immunoglobulin allotype with anti-polysaccharide antibodies in Caucasian adults.

Km allotype antigens are serologic markers expressed on kappa light chains of human immunoglobulins. To determine whether th Km phenotype of an individual is related to his ability to make antibodies to polysaccharide antigens, we correlated the Km allotypes of 129 healthy caucasian adults with the concentrations of specific antibodies to three bacterial polysaccharide antigens after immunization. The 14 individuals expressing the Km(1) allotype had lower concentrations of IgG, IgM, and IgA antibodies by enzyme-linked immunosorbent assay and total antibody by radioimmunoassay to Haemophilus influenzae type b and Neisseria meningitidis group C capsular polysaccharides when compared with the 115 Km(1) negative individuals. The Km-associated differences in H. influenzae type b and N. meningitidis group C antibody concentrations were confined to kappa light chain-containing antibody (P = 0.029 and 0.003, respectively). Similarly, the Km(1) positives had slightly lower kappa chain-containing Ig than the Km(1) negatives (P = 0.079). We conclude that genes in or near the kappa light chain locus play a role in the regulation of kappa-containing antibody production to some bacterial polysaccharides and perhaps to other antigens.

Correlation between G2m(n) immunoglobulin allotype and human antibody response and susceptibility to polysaccharide encapsulated bacteria.

To determine whether genetic factors influence the human antibody response to polysaccharides, we correlated Ig allotypes with the concentrations of antibody to 14 bacterial capsular antigens in 130 actively immunized Caucasian adults. The 88 individuals possessing G2m(n), an allotype antigen of IgG2 subclass heavy chains, had significantly higher postimmunization antibody levels to Haemophilus influenzae type b (Hib) and 8 of 11 pneumococcal types (P less than 0.05) than the 42 lacking this antigen. For Hib, pneumococcus type 14, and meningococcus group C, an increased response was observed in IgG class but not in IgM or IgA classes of antibody. The G2m(n) positive individuals also had higher preimmunization antibody levels to most polysaccharide antigens. Total IgG2 concentrations were correlated with the mean postimmunization antibody concentrations to pneumococci (P = 0.005), but this correlation was independent of G2m(n) by multiple regression analysis. To determine if the lack of G2m(n) was associated with increased susceptibility to infection, we compared the frequencies of various Ig allotypes in 98 children infected with Hib and 98 matched controls. Caucasian children with Hib infections other than epiglottitis were significantly more likely to lack the G2m(n) allotype than controls (P less than 0.05). G2m(n) negative Caucasian children less than or equal to 18 mo old have a 5.1-fold higher risk of nonepiglottitic Hib infections than G2m(n) positive children (P less than 0.01). We conclude that allotypic variants of the gamma-2 heavy chain genes, or genes in linkage equilibrium with them, exert a regulatory influence on the caucasian antibody response to a variety of immunologically distinct bacterial polysaccharide antigens. Young Caucasian children of the low responder phenotype, i.e., those lacking the G2m(n) allotype, are genetically predisposed to Hib and perhaps other bacterial infections.

Proliferation of resting B cells is modulated by CR2 and CR1.

Absence of the third component of complement, C3, is associated with impaired ability to synthesize antibody, particularly in the presence of limiting antigen [1-9]. The mature B lymphocyte bearing the surface immunoglobulin receptor transduces signals for proliferation and differentiation upon binding of specific antigen. This mature B cell also bears two related membrane proteins, CR2 (the C3d/Epstein-Barr virus receptor) (CD35) [15], which can mediate the binding of ligands to which appropriate cleavage fragments of C3 have become attached [16]. It has been suggested that these receptors play a direct role(s) in B cell activation [17-25]. In light of previous in vivo observations we decided to assess the function of CR2 and CR1 in relation to B cell activation through the membrane IgM receptor. Highly purified splenic B cells were prepared. No contaminating T cells or macrophages were detected by flow cytometric analysis and no proliferative activity was present upon PHA or ConA stimulation of the purified cells. The B cells were separated into low (activated), medium (preactivated) and high density (resting) fractions by Percoll gradient density centrifugation [26]. The responses of the B cell subpopulations to various concentrations of anti mu (DA4.4 monoclonal antibody) [27] were examined for proliferation at 72 h and for IgM/IgG production at 7 days. Low density B cells were maximally stimulated and no concentration of anti-mu was effective in enhancing their responses. High density B cells proliferated to anti-mu in a concentration dependent manner. When substimulatory concentrations of anti-mu were employed, concomitant crosslinking of CR2 (with either of 2 distinct monoclonal antibodies HB-5 [28] or OKB7 [17]) resulted in a 45% enhancement of B cell proliferation above that observed by crosslinking of SIgM alone. In these studies, total IgM and IgG did not increase in the absence of T cells or T cell factors, indicating that terminal differentiation did not occur. In contrast, when a monoclonal antibody to CR1(44D) [29] was employed in an identical experiment, B cell proliferation was completely inhibited. Antibodies to CR2 or CR1 either alone or in crosslinked form did not enhance B cell proliferation. Immune complexes may crosslink the B cell surface in a manner analogous to our model when the immunoglobulin receptor and CR2 are simultaneously engaged. This activation signal may be particularly important in eliciting antibody responses when the quantity of specific antigen or the affinity for antigen is low. The marked inhibition of proliferation induced by CR1 suggests an alternate role for this receptor in modulation of B cell responses.

IgG1, IgG2 and IgM responses to two Haemophilus influenzae type b conjugate vaccines in young infants.

PRP-meningococcal outer membrane protein complex (PRP-OMPC) and oligosaccharide linked to variant diphtheria toxin (HbOC) Haemophilus influenzae type b (HIB) conjugate vaccines have both been licensed for United States infants at 2 months of age. Differences in serologic responses for these vaccines have been noted with PRP-OMPC producing an early response at 2 months of age and HbOC producing a higher response after a third dose at 6 months of age. To further characterize the nature of these distinct responses, we measured the IgG1, IgG2 and IgM anti-HIB concentrations by enzyme-linked immunosorbent assay after administration of both vaccines. PRP-OMPC produced an IgM and IgG1 anti-HIB response following the initial dose at 2 months of age. After two doses of HbOC an increase in IgG1 and IgM were noted and after a third dose at 6 months of age an IgG2 anti-HIB response occurred. In addition 33 study subjects were boosted with PRP-OMPC at age 18 months and compared with 34 subjects who received only a primary dose. The anti-HIB IgG1 and IgG2 concentrations following the booster dose were both significantly higher for the primed group (P = 0.0001 and P = 0.001, respectively). Both HIB conjugate vaccines produce predominantly IgG1 anti-HIB antibody responses. The early response to PRP-OMPC vaccine at 2 months of age may result from adjuvant characteristics of the OMPC.

Enzyme-linked immunosorbent assay to assess respiratory syncytial virus concentration and correlate results with inflammatory mediators in tracheal secretions.

OBJECTIVE: We developed an enzyme-linked immunosorbent assay (ELISA) for the quantitation of respiratory syncytial virus (RSV) in respiratory secretions in intubated patients infected with RSV. METHODS: We compared the quantitative ELISA and a standardized plaque assay in intubated children <2 years of age who were mechanically ventilated for severe RSV disease and enrolled in a randomized double blind placebo-controlled treatment trial of a monoclonal antibody to the F protein of RSV (palivizumab; Synagis). We also examined the relationship between the concentrations of virus as measured by ELISA and of three inflammatory indices in respiratory secretions (white blood cell count, myeloperoxidase and eosinophilic cationic protein). RESULTS: Quantitative ELISA and plaque assay were highly correlated for both tracheal aspirates (r = 0.67, P = 0.001) and nasal wash specimens (r = 0.75, P = 0.001). Treatment with palivizumab significantly neutralized RSV in tracheal aspirates as measured by plaque assay. In contrast quantitation of RSV by ELISA was not affected by palivizumab treatment. This finding is consistent with results that were obtained in preliminary studies of RSV-containing media treated with monoclonal antibody, where we found that the ELISA measured virus whether antibody-bound or not. The inflammatory indices were not correlated with RSV concentration measured by ELISA or plaque assay. CONCLUSIONS: We conclude that this quantitative ELISA is a potentially useful tool for measurement of RSV concentration in respiratory secretions that may help elucidate the pathophysiology of acute RSV infection. Specific antiviral strategies for the treatment of RSV disease could be evaluated by this method.

Impaired polysaccharide responses in immunodeficient patients: relevance to bone marrow transplant patients.

Bone marrow transplant patients are at increased risk for pneumococcal and H. influenzae type b (HIB) infections. These polysaccharide encapsulated bacteria are also pathogens for young healthy children. In healthy children age related susceptibility has been associated with poor response to polysaccharides and low serum IgG2 subclass antibody concentrations. Of note, immune reconstitution following bone marrow transplantation has been characterized by slow return of both the response to polysaccharides and IgG2 serum concentrations. Thus immune reconstitution following bone marrow transplantation is similar to the maturation that occurs in healthy children. In addition to transplant patients, we have identified five groups of immunodeficient patients who are at increased risk for polysaccharide encapsulated pathogens. IgG2 subclass deficient patients were shown to have impaired responses to polysaccharide antigens. We have also defined "Selective Antibody Deficiencies" which are individuals with normal IgG subclass concentrations but poor responses to polysaccharide antigens. Next, patients who developed HIB disease in spite of immunization (i.e. HIB vaccine failures) were demonstrated to have lower serum IgG2 and IgG4 subclass concentrations compared to controls. Finally, Native Americans (an ethnic group with an increased incidence of pneumococcal and HIB infections) were shown to be poor responders to polysaccharide antigens and have significantly lower concentrations of serum IgG2 and IgG4 as compared to controls. New HIB polysaccharide vaccines linked to protein antigens (conjugate vaccines) have been developed that are more immunogenic in healthy young children. In addition, these conjugate vaccines have resulted in protective responses in each of the newly described immunodeficient groups. We therefore now propose to evaluate bone marrow transplant patients' response to HIB conjugate vaccine.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of multiple immunization schedules for Haemophilus influenzae type b-conjugate and tetanus toxoid vaccines following bone marrow transplantation.

Antibody concentrations to vaccine-preventable diseases decline following BMT and an optimal schedule for vaccination after transplant has not been established. We examined antibody responses to tetanus toxoid (TT) and Haemophilus influenzae type b-conjugate (HIB) vaccines of BMT patients immunized at 6, 12 and 24 months (6 month group, n = 21) and compared them to those previously reported for patients immunized at 3, 6, 12 and 24 months (3 month group, n = 74) or at 12 and 24 months (12 month group, n = 17) following transplantation. Geometric mean total anti-HIB and IgG anti-TT concentrations were significantly higher after the 12 month dose in the 3 and 6 month immunization groups compared to the group who received their first dose at 12 months. Although HIB antibody concentrations were higher in the 3 month and 6 month groups 12 to 24 months after BMT, the proportion of patients with protective levels was not significantly different from the proportion protected in the 12 month group. Following the 24 month immunizations, geometric mean antibody concentrations to HIB and TT were similar for all three immunization groups. The proportion of patients in each group with protective levels of HIB antibody after the 24 month dose was > or = 80%. A two dose schedule of HIB and TT vaccines at 12 and 24 months after BMT should afford protection.

Haemophilus influenzae type b (HIB)-conjugate immunization before bone marrow harvest in autologous bone marrow transplantation.

Immune reconstitution following autologous bone marrow transplantation (ABMT) is characterized by defects in B cell and T cell function and loss of specific antibody. In the late post-transplant period, patients are at risk for infections with polysaccharide encapsulated organisms and respond poorly to polysaccharide vaccines. We examined whether immunizing ABMT patients before bone marrow (BM) harvest enhanced the early recovery of specific antibody. Twelve patients were immunized before BM harvest with Haemophilus influenzae type b (HIB)-conjugate, tetanus toxoid and polysaccharide pneumococcal vaccines. Forty-one comparable ABMT patients not immunized prior to BM harvest were also studied. Following ABMT, both groups of patients were immunized with HIB-conjugate and tetanus toxoid vaccines at 3, 6, 12 and 24 months and with pneumococcal vaccine at 12 and 24 months. Patients immunized before BM harvest had higher HIB antibody concentrations during the first 2 years post-transplant, the differences reaching significance at 3 months (P = 0.0001) and following the 24-month dose (P = 0.048). Tetanus toxoid antibody concentrations were also significantly higher at 3 months (P = 0.001) and 6 months (P = 0.032) in patients immunized before BM harvest. There were no differences in pneumococcal antibody concentrations between the two groups. Immunization of patients before bone marrow harvest resulted in higher anti-HIB antibody concentrations following ABMT and may be an effective strategy to prevent infectious complications.

Antibody responses to tetanus toxoid and Haemophilus influenzae type b conjugate vaccines following autologous peripheral blood stem cell transplantation (PBSCT).

Accelerated granulocyte and platelet recovery following peripheral blood stem cell transplantation (PBSCT) are well documented. We hypothesize that functional immunity may also be enhanced in PBSCT and performed a phase II trial of immunizations in patients with lymphoma undergoing autologous transplantation with peripheral blood stem cells or bone marrow. Seventeen BMT and 10 PBSCT recipients were immunized at 3, 6, 12, and 24-months post-transplantation with Haemophilus influenzae type b (HIB)-conjugate and tetanus toxoid (TT) vaccines. IgG anti-HIB and anti-TT antibody concentrations were measured and compared between the two groups. Geometric mean IgG anti-HIB antibody concentrations were significantly higher for PBSCT recipients compared to BMT recipients at 24 months post-transplantation (11.3 micrograms/ml vs 0.93 microgram/ml, P = 0.051) and following the 24 month immunization (66.2 micrograms/ml vs 1.30 micrograms/ml, P = 0.006). Similar results were noted for IgG anti-TT antibody with significantly higher geometric mean antibody concentrations in the PBSCT group at 24 months post-transplantation (182 micrograms/ml vs 21.6 micrograms/ml, P = 0.039). Protective levels of total anti-HIB antibody were achieved earlier in PBSCT recipients compared with those of BMT recipients. PBSCT recipients had higher antigen-specific antibody concentrations following HIB and TT immunizations. These results suggest enhanced recovery of humoral immunity in PBSCT recipients and earlier protection against HIB with immunization.

Critical appraisal of immunization strategies for prevention of infection in the compromised host.

Chemotherapy and/or bone marrow transplantation clearly places individuals at increased risk for many vaccine-preventable diseases. Both active and passive immunization strategies have been considered for protecting immunocompromised patients. Many compromised hosts are capable of mounting protective responses to some protein toxoids, polysaccharide-conjugate, and viral vaccines. Effective, safe, and practical dosage schedules are evolving for individual vaccines and specific patient populations.

Response to Haemophilus influenzae type B conjugate vaccine in children undergoing splenectomy.

Asplenic children are at increased risk for serious infection with polysaccharide encapsulated bacteria including Haemophilus influenzae type b (HIB), Streptococcus pneumoniae, and Neisseria meningitidis. Immunization with polysaccharide vaccines is recommended for children undergoing splenectomy. In 1987 a new more immunogenic HIB vaccine was licensed in the US to replace the pure HIB polysaccharide vaccine that was licensed in 1985. This polysaccharide-conjugate vaccine consists of the HIB polysaccharide linked to a protein carrier, diphtheria toxoid. Therefore, we evaluated the immune response of children undergoing splenectomy to HIB-conjugate vaccine. Thirteen children (7 with Hodgkin's disease, 4 with idiopathic thrombocytopenia, 2 with hereditary spherocytosis) aged 3 to 19 years were immunized with HIB-conjugate vaccine prior to splenectomy and serum was obtained following splenectomy. In addition, 15 healthy control children aged 2 to 14 years were immunized with the pure polysaccharide HIB vaccine for comparison. The patients undergoing splenectomy who received the HIB-conjugate vaccine had a geometric mean IgG anti-HIB antibody concentration of 48,106 ng/mL versus 10,786 ng/mL for the control patients who received the pure polysaccharide vaccine (P = .01). The presumed protective level of antibody is 1,000 ng/mL and all children were well above this concentration. Therefore, we propose that children undergoing splenectomy be immunized with an HIB-conjugate vaccine.

Human polysaccharide-specific B cells are responsive to pokeweed mitogen and IL-6.

The responsiveness of polysaccharide-specific B cells to PWM was examined in vitro. Spleen cells from six patients immunized with Haemophilus influenzae type b-diphtheria toxoid, pneumococcal and meningococcal vaccines were T cell-depleted and separated by Percoll density gradient centrifugation. In each B cell fraction, spontaneous antibody production was demonstrated to capsular polysaccharides as well as diphtheria toxoid. The peak of spontaneous antibody production was demonstrated to be five to seven days after immunization. When T cells and PWM were added, the total Ig secretion increased in all B cell fractions. PWM also enhanced IgG antibody directed to each of three polysaccharide Ag measured. This enhancement was most noticeable for nonresting B cells. The PWM effect was not confined to IgG, as IgM and IgA to Neisseria meningitidis type C were measured and also enhanced. The kinetics of the PWM response demonstrated the most IgG antibody to polysaccharide Ag from spleens immunized five to seven days before splenectomy. When the patients were immunized either 2 days or 4 mo before splenectomy, no spontaneous IgG antibody to polysaccharides was detected although PWM induced small amounts of antibody. Finally, anti-IL-6 antibody blocked PWM-induced total and polysaccharide-specific antibody production. We conclude that human polysaccharide-specific B cells are responsive to PWM and IL-6. We suggest that polysaccharide B cells are not truly "T cell-independent" and may respond to T cell lymphokines and thus are similar to protein-specific B cells.

Blood-group-A-like substance in a preparation of pneumococcal vaccine.

A platelet transfusion from a blood group O donor, immunized 1 month before with Pneumovax, caused a hemolytic reaction in a blood group A recipient. Forty-five of 59 group O donors (76%) and all of nine group B donors immunized with Pneumovax had a fourfold or higher anti-A response. Half of the anti-A antibody in high titered donors was in the IgG fraction. Pneumovax contained approximately 30 micrograms of an A-like substance per dose; polyvalent pneumococcal vaccines prepared by two other manufactures contained very low and probably subimmunogenic concentrations. Several culture media prepared from animal tissues contained as antigen of similar physical, immunologic, and chemical properties, and were the most likely source of the contaminant. Manufacturing procedures have since been revised to eliminate A-like substances.

Haemophilus influenzae type b unsuspected bacteremia.

To further define the clinical features and natural history of unsuspected Haemophilus influenzae type b (Hib) bacteremia, we retrospectively reviewed the records of 322 Hib infections observed during a 45-month period at Children's Hospital, Boston. We identified 31 patients with unsuspected Hib bacteremia and 19 with unsuspected Hib antigenemia and sterile blood cultures. Bacteremic patients were typically under two years of age (81%), had high fevers (mean = 39.5 degrees C), and frequently had otitis media (65%) diagnosed as their only focus of infection at presentation. Nineteen of 31 were empirically treated with oral antibiotics. Ten of 31 (32%) developed focal infectious complications consisting of meningitis (n = 7), cellulitis (n = 2), and pneumonia (n = 1). Children with focal infectious complications differed from those without infectious complications in having a significantly higher mean fever of 40.3 degrees C compared to 39.7 degrees C (P less than 0.05). Five of 19 (26%) empirically treated patients developed focal complications (all meningitis) compared to five of 12 (42%) untreated patients. Blood cultures at follow-up visit were positive in three of 19 (9%) treated patients and seven of 12 (42%) untreated patients (P less than 0.05). Of the 19 children with antigenemia and sterile blood cultures, 16 (84%) were empirically treated with antibiotics, and none had positive blood cultures or focal infections on follow-up evaluation. Children with occult Hib bacteremia are at high risk for developing serious focal infections, particularly meningitis, despite empiric antibiotic therapy. Once Hib bacteremia is suspected, strong consideration should be given to parenteral in hospital antibiotic therapy. The utility of rapid antigen detection for identifying high-risk patients requires further evaluation.

Passive immunization against disease due to Haemophilus influenzae type b: concentrations of antibody to capsular polysaccharide in high-risk children.

From the pooled plasma of 54 adult donors immunized with Haemophilus influenzae type b capsular polysaccharide (CP), a human hyperimmune globulin termed bacterial polysaccharide immune globulin (BPIG) was prepared. The pharmacokinetics of antibody to H. influenzae type b CP in high-risk children was compared after BPIG and conventional immune serum globulin (ISG) were administered intramuscularly at a dose of 0.5-0.6 ml/kg. The increase in antibody level four to seven days after injection was ninefold higher with BPIG than with ISG and remained higher throughout the three-month follow-up period. The mean half-life of antibody to H. influenzae type b CP was similar after BPIG (27 days) and ISG (29 days). Antibody levels returned to baseline one to two months after ISG but remained significantly above baseline three months after BPIG (mean concentration, 385 ng/ml). Based on the assumption that 150 ng of IgG antibody to H. influenzae type b CP/ml is the minimal protective level, it is concluded that a single intramuscular dose of 0.5 ml of BPIG/kg will protect children for four months.

Kappa and lambda light chain composition of antibody to the capsular polysaccharide of Haemophilus influenzae type b.

Concentrations of total immunoglobulins bearing kappa and lambda light chains were measured in the sera of 215 healthy white children aged 6 months to 10 years. Both kappa and lambda concentrations increased with age. However, the concentration of immunoglobulins bearing kappa light chains increased at a greater rate than those bearing lambda light chains (P = .01). Thus, the kappa:lambda ratio of the youngest children (6-24 months) was significantly lower than that of the oldest (25-130 months) (P = .0015). The relationship between the IgG antibody concentration and the light chain composition of the specific antibody directed to Haemophilus influenzae type b (Hib) polysaccharide was also assessed in 62 of 215 serum samples with detectable Hib antibody. The IgG Hib antibody concentration was strongly correlated with the kappa:lambda Hib antibody ratio (r = .60, P = .001), and this correlation was independent of age. Thus, light chain selection and response to polysaccharides may be regulated by common mechanisms that mature late in ontogeny.

Recurrent sinopulmonary infection and impaired antibody response to bacterial capsular polysaccharide antigen in children with selective IgG-subclass deficiency.

We studied 20 children with recurrent sinopulmonary infections and serum IgG levels within the normal range, who had selective IgG-subclass deficiency. Twelve of the children were IgG2 deficient, five were IgG3 deficient, and three were deficient in both IgG2 and IgG3. IgA deficiency was present in 3 of the 20 patients. In the children with IgG2 deficiency, serum antibody concentrations to the capsular polysaccharide of Hemophilus influenzae type B (Hib) were significantly lower than those in age-matched controls, both before and after immunization with the Hib capsular polysaccharide antigen, which elicits antibody predominantly of the IgG2 subclass. In contrast, their serum antibody titers to the tetanus and diphtheria toxoid protein antigens, which elicit antibody predominantly of the IgG1 subclass, were normal in comparison with those of age-matched controls. These results suggest that impairment of the antibody response to specific microbial antigens predisposes patients with selective IgG-subclass deficiencies to recurrent infections. Thus, as an aid in determining therapy, children with recurrent infections and normal total serum IgG should be evaluated for this condition.

Human B cells secrete predominantly lambda L chains in the absence of H chain expression.

Ig H and L chains are independently assembled in B cells and then secreted together as a functional protein. H chains cannot be secreted without assembly to L chains; however, L chains can be secreted in the absence of H chains by both mice and human cells. To examine the influence of H chain expression on human L chain isotype selection (kappa or lambda), we compared the kappa/lambda ratio of L chains unassociated with H chains (free L chains) to the kappa/lambda ratio of L chains associated with H chains. Culture supernatants of human splenocytes were assayed for kappa and lambda L chains. Free L chains were the predominant form of L chains detected in unstimulated cultures, accounting for 68 to 70% of the total. This was in contrast to the minor proportion that free L chains represented (less than 20%) in cultures stimulated with PWM or LPS (p less than 0.01). Furthermore, the kappa/lambda ratio of light chains detected in unstimulated cultures was 0.5 as compared to 1.3 for PWM stimulated cultures (p = 0.0001). To demonstrate that the decreased kappa/lambda ratio of L chains in the supernatants of cultures of unstimulated B cells was due to free L chains, we measured the kappa/lambda ratio of IgG and IgM-associated L chains. In both the stimulated and unstimulated cultures, the kappa/lambda ratio of L chains associated with H chains was greater than the ratio determined for free L chains. Free L chains were shown to be predominantly lambda as compared to the predominantly kappa phenotype of L chains associated with H chains. Thus absence of H chain expression affects selection of L chain isotypes secreted by human B cells.