Maternal IgA2 Recognizes Similar Fractions of Colostrum and Fecal Neonatal Microbiota.

Microbiota acquired during labor and through the first days of life contributes to the newborn's immune maturation and development. Mother provides probiotics and prebiotics factors through colostrum and maternal milk to shape the first neonatal microbiota. Previous works have reported that immunoglobulin A (IgA) secreted in colostrum is coating a fraction of maternal microbiota. Thus, to better characterize this IgA-microbiota association, we used flow cytometry coupled with 16S rRNA gene sequencing (IgA-Seq) in human colostrum and neonatal feces. We identified IgA bound bacteria (IgA+) and characterized their diversity and composition shared in colostrum fractions and neonatal fecal bacteria. We found that IgA2 is mainly associated with Bifidobacterium, Pseudomonas, Lactobacillus, and Paracoccus, among other genera shared in colostrum and neonatal fecal samples. We found that metabolic pathways related to epithelial adhesion and carbohydrate consumption are enriched within the IgA2+ fecal microbiota. The association of IgA2 with specific bacteria could be explained because these antibodies recognize common antigens expressed on the surface of these bacterial genera. Our data suggest a preferential targeting of commensal bacteria by IgA2, revealing a possible function of maternal IgA2 in the shaping of the fecal microbial composition in the neonate during the first days of life.

ANTIGENIC STIMULATION DURING PREGNANCY MODIFIES SPECIFIC IGA1 AND IGA2 SUBCLASSES IN HUMAN COLOSTRUM ACCORDING TO THE CHEMICAL COMPOSITION OF THE ANTIGEN.

BACKGROUND: Several studies have evaluated the effect of infectious diseases and vaccine protocols during pregnancy on maternal milk immunoglobulin (Ig) levels, to understand the protection conferred by lactation on newborns. Colostrum is the primary source of maternal IgA for the newborn. IgA participates in protection mechanisms in the neonate's mucosa. In humans, IgA has two subclasses with differential anatomical distribution among mucosal compartments. Total IgA levels in maternal milk vary after antigen stimulation and have differential affinities in function of the chemical composition of the antigens. We studied the effect of antigenic stimulation during pregnancy on the concentrations of specific IgA1 and IgA2 subclasses in human colostrum. METHODS: We analyzed data from 113 women in Mexico City and compared the amount of IgA subclasses in colostrum against three antigens: two from vaccine protocols (tetanus toxoid and pneumococcal polysaccharides) and lipopolysaccharide, a ubiquitous antigen in the gastrointestinal tract. RESULTS: In agreement with the previous reports, we showed that IgA1 from colostrum mainly recognized protein antigens; in sharp contrast, IgA2 was mostly directed against polysaccharide antigens. These levels increased in women who had previous contacts through vaccination or infections during pregnancy. CONCLUSIONS: Antigen interaction during pregnancy increased the amount of specific IgA subclasses, depending on the chemical composition of the antigen.

The Profile of Immunoglobulin A and Immunoglobulin G Subclasses in Sprague Dawley Rats Experimentally Infected with Brucella abortus Biotype 1.

This study measured total serum immunoglobulin A (IgA), immunoglobulin G (IgG)1, IgG2a response against whole cell antigen (WCA), outer membrane protein (OMP), periplasmic protein (PP), cytoplasmic protein (CP), and crude Brucella protein (CBP) of Brucella abortus in experimental brucellosis induced with B. abortus biotype 1 in Sprague Dawley (SD) rats during a 17-week infection period. Six- to 8-week-old SD rats (n = 44) were experimentally infected with 1 × 109 colony forming unit of B. abortus biotype 1 through the intraperitoneal route. Serial serum samples were collected from the rat at 0, 3, 7, 14, 21, 28, 35, 42, 60, 90, and 120 days after inoculation. The sera were tested by enzyme linked immunosorbent assay. We have noticed a very low level and short persistence of IgA antibody in our experiment. The low level and short persistence of IgA antibody suggest that this antibody isotype might not be protective against brucellosis in rats. Both Th1 and Th2 specific immune responses were recorded in our study with the production of IgG1 and IgG2a antibody isotopes, respectively. We noticed significant dominant IgG2a antibody responses over IgG1 responses throughout the experiment (p < 0.001) against WCA and OMP. The mixed Th1 and Th2 dominant immune responses mediated by IgG2a and IgG1 antibody isotypes were observed against CP, PP, and CBP. Data of our study suggest that IgG2a dominant responses in the early stages of disease play the main role in conferring protection against brucellosis and with the progress of disease IgG1 dominant responses were elicited.

Assignment of Serotype-Specific IgG1, IgG2, and IgA Weight-Based Antibody Units to the Human Pneumococcal Standard Reference Serum, 007sp.

In 2011, the human pneumococcal standard reference serum, 007sp, was established as a replacement for the previous standard, lot 89SF, supplies of which were dwindling. The pneumococcal reference serum is used primarily in the standardized pneumococcal enzyme-linked immunosorbent assay (World Health Organization reference enzyme-linked immunosorbent assay) but has also been used in functional assays. Serotype-specific IgG values for 24 pneumococcal capsular serotypes have previously been assigned to 007sp by bridging to the original values derived for lot 89SF. In this study, by bridging to existing values in lot 89SF, we assign weight-based serotype-specific IgA, IgG1, and IgG2 to 007sp for 11 pneumococcal capsular serotypes (1, 3, 4, 5, 6B, 7F, 9V, 14, 18C, 19F, and 23F), as well as serotype 19A-specific IgA. Concentrations for serotype-specific IgA, IgG1, and IgG2 present in 007sp were comparable to those previously assigned to lot 89SF. In addition, the concentration of serotype-specific IgG1 plus IgG2 assigned to 007sp significantly correlated to previously assigned 007sp IgG values. The accuracy of antibody assignments to 007sp from lot 89SF was assessed by comparing the concentration of serotype-specific IgA, IgG1, and IgG2 in 16 unknown samples using both 007sp and lot 89SF as the standard. Interpolated values for the unknown samples were highly correlated with average R2 values of 0.9729, 0.9951, and 0.9933 for IgA, IgG1, and IgG2, respectively, for all serotypes demonstrating the precise nature assignments to 007sp made in this study. Nonparallelism between 007sp and lot 89SF has precluded the derivation of serotype-specific IgM values.IMPORTANCE A well-characterized antibody standard is an indispensable reagent for use in assays designed to measure antibodies with precision and where assays between laboratories need to be comparable. The human pneumococcal standard reference serum, lot 89SF, greatly facilitated the standardization of enzyme-linked immunosorbent assay methodologies during a critical period when the first pneumococcal polysaccharide-conjugate vaccines were being evaluated for licensure. Due to dwindling supplies of lot 89SF, a new reference standard, 007sp, was produced in 2011. Understanding the isotype and subclass composition of either natural or vaccine induced responses to pathogens has assumed increasing importance. In this study, we have assigned IgA, IgG1, and IgG2 values to pneumococcal serotypes 1, 3, 4, 5, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F by bridging to existing values in lot 89SF.

Exploring Site-Specific N-Glycosylation of HEK293 and Plant-Produced Human IgA Isotypes.

The full potential of recombinant Immunoglobulin A as therapeutic antibody is not fully explored, owing to the fact that structure-function relationships of these extensively glycosylated proteins are not well understood. Here monomeric IgA1, IgA2m(1), and IgA2m(2) variants of the anti-HER2 antibody (IgG1) trastuzumab were expressed in glyco-engineered Nicotiana benthamiana plants and in human HEK293-6E cells. All three IgA isotypes were purified and subjected to biophysical and biochemical characterization. While no differences in assembly, antigen binding, and glycosylation occupancy were observed, both systems vary tremendously in terms of glycan structures and heterogeneity of glycosylation. Mass-spectrometric analysis of site-specific glycosylation revealed that plant-produced IgAs carry mainly complex-type biantennary N-glycans. HEK293-6E-produced IgAs, on the contrary, showed very heterogeneous N-glycans with high levels of sialylation, core-fucose, and the presence of branched structures. The site-specific analysis revealed major differences between the individual N-glycosylation sites of each IgA subtype. Moreover, the proline-rich hinge region from HEK293-6E cell-derived IgA1 was occupied with mucin-type O-glycans, whereas IgA1 from N. benthamiana displayed numerous plant-specific modifications. Interestingly, a shift in unfolding of the CH2 domain of plant-produced IgA toward lower temperatures can be observed with differential scanning calorimetry, suggesting that distinct glycoforms affect the thermal stability of IgAs.

[Relationship between M-Protein of Multiple Myeloma and False Positive Syphilis Serological Results].

BACKGROUND: The false positive in conventional syphilis serological test was found in patients with multiple myeloma (MM). OBJECTIVE: To investigate the relationship between the M-protein of patients with MM and the false positive in conventional syphilis serologic test. METHODS: The M-protein of 68 MM cases was typed with immunofixation electrophoresis and 68 cases of MM were screened with non-specific and specific syphilis serologic tests, then the samples with syphilic serological positive were chosen and confirmed with immonobloting test, finally the relationship between M protein of MM and the false positive of syphilis serological test were analysed. RESULTS: Four out of 68 cases showed the positive in syphilis serological test and further were confimed to be false positive by immunoblotting test, the false positive rate was nearly 6%. The M-protein of MM patients in our hospital mostly possessed IgG, κ type, followed by IgA, κ type, light chain κ type. In general, κ : λ = 2.4 : 1. Among samples of 4 cases with syphilis serological positive 2 cases were of IgG and κ type, 1 case was of IgG, λ type, another 1 case was IgA, κ type. CONCLUSION: The M-protein of IgG and IgA types in MM patients results in syphilis serological false positive reaction. The clinicians and laboratorial technicians should pay a great attention to screen the MM patients for the false positive syphilis serological test so as to avoid the misdiagnosis and subsequent embarassment.

Comparison of different ELISA protocols for the detection of IgA against influenza nucleoproteins in trachea of vaccinated chickens.

Vaccines targeting mucosal immunity are important for the control of infection by pathogens with mucosal portals of entry, such as avian influenza. However, reliable and effective methods for determining levels of mucosal IgA stimulated by vaccination are not well developed in poultry and are necessary for determining efficacy. The objective of the present study was to compare different ELISA protocols to evaluate levels of mucosal IgA against two different sequences of nucleoprotein (NP:), a highly conserved internal protein in avian influenza virus, in trachea. Positive control tracheas were obtained through hyperimmunization of birds with adjuvated NP1 and NP2 peptide conjugated with keyhole limpet hemocyanin administered both orally and parenterally; negative birds received no antigen. Trachea samples were homogenized, and supernatant fluid was collected to separate IgA. ELISA was performed on NP1- or NP2-positive trachea samples, negative trachea samples, and blank wells with different levels of NP1 and NP2 coating peptides (5 or 10 µg/mL) using two different secondary antibodies (Gene Tex, GT:, or Thermo Scientific, TS:), with or without an acetate wash, and using maximum, medium, or low binding ELISA plates. The TS antibody resulted in a higher background signal compared to GT. Furthermore, coating plate wells with NP2 resulted in very high background compared to NP1. An acetate buffer wash resulted in the muffling of signals, and medium and low binding plates used in the study resulted in better results than maximum binding plates. These results suggest that the selection of appropriate secondary antibodies, binding plates, and ELISA reagent protocols all play important roles in determining NP1- or NP2-specific IgA levels in trachea samples.

Identification of distinct glycoforms of IgA1 in plasma from patients with immunoglobulin A (IgA) nephropathy and healthy individuals.

Immunoglobulin A nephropathy (IgAN) is the most common form of glomerulonephritis worldwide and is histologically characterized by the deposition of IgA1 and consequent inflammation in the glomerular mesangium. Prior studies suggested that serum IgA1 from IgAN patients contains aberrant, undergalactosylated O-glycans, for example, Tn antigen and its sialylated version, SialylTn (STn), but the mechanisms underlying aberrant O-glycosylation are not well understood. Here we have used serial lectin separation technologies, Western blot, enzymatic modifications, and mass spectrometry to explore whether there are different glycoforms of IgA1 in plasma from patients with IgAN and healthy individuals. Although total plasma IgA in IgAN patients was elevated ∼ 1.6-fold compared with that in healthy donors, IgA1 in all samples was unexpectedly separable into two distinct glycoforms: one with core 1 based O-glycans, and the other exclusively containing Tn/STn structures. Importantly, Tn antigen present on IgA1 from IgAN patients and controls was convertible into the core 1 structure in vitro by recombinant T-synthase. Our results demonstrate that undergalactosylation of O-glycans in IgA1 is not restricted to IgAN and suggest that in vivo inefficiency of T-synthase toward IgA1 in a subpopulation of B or plasma cells, as well as overall elevation of IgA, may contribute to IgAN pathogenesis.

Comprehensive endometrial immunoglobulin subclass analysis in infertile women suffering from repeated implantation failure with or without chronic endometritis.

PROBLEM: Chronic endometritis (CE) is a local inflammatory condition with unusual plasmacyte infiltration in the endometrial stromal area. CE is frequently found in infertile women with repeated implantation failure (RIF). In this study, we comprehensively investigated the endometrial immunoglobulin (Ig) subclass expression in infertile women suffering from RIF with versus without CE. METHOD OF STUDY: Endometrial biopsy specimens obtained from 28 infertile women with RIF and CE (the RIF-CE group), 23 infertile women with RIF but without CE (the RIF-non-CE group), and 22 proven fertile women undergoing hysterectomy for benign endometrial pathology (the control group) were immunostained for Ig subclass expression. RESULTS: The density of IgM+, IgA1+, IgA2+, IgG1+, and IgG2+ stromal cells were significantly higher in the RIF-CE group than that in the RIF-non-CE and control group. The density of IgG2+ stromal cells was significantly higher than that of any other Ig subclass-positive cells (P<0.045) in the RIF-CE group. In serial section staining, the immunoreactivity for CD138 and Ig subclasses in the endometrial stroma was detectable in adjacent cells of some specimens in the RIF-CE group. CONCLUSIONS: The endometrium of infertile women with RIF-CE was characterized by increase in IgM, IgA, and IgG expression and predominance of IgG2 over other Ig subclasses.

High prevalence of IgA class anti-neutrophil cytoplasmic antibodies (ANCA) is associated with increased risk of bacterial infection in patients with cirrhosis.

BACKGROUND & AIMS: Anti-neutrophil cytoplasmic antibodies (ANCA) are a non-uniform family of antibodies recognizing diverse components of neutrophil granulocytes. ANCA formation might be induced by protracted bacterial infections or probably reflect an abnormal immune response to commensal microorganisms. Bacterial infections are common complications in cirrhosis with high incidence of episodes caused by enteric organisms, therefore, we sought to study the presence and clinical importance of ANCA in cirrhosis. METHODS: Sera of 385 patients with cirrhosis of different etiologies were assayed for ANCA of IgG, IgA, IgA1, IgA2, and secretory IgA subtypes by indirect immunofluorescence and ELISAs. The control group comprised 202 patients with chronic liver diseases without cirrhosis and 100 healthy subjects. In cirrhosis, a 2-year follow-up, observational study was conducted to assess a possible association between the presence of ANCA and clinically significant bacterial infections. RESULTS: Prevalence of ANCA IgA was significantly higher in cirrhosis (52.2%) compared to chronic liver diseases (18.6%) or healthy controls (0%, p<0.001 for both). ANCA IgA subtyping assays revealed marked increase in the proportion of IgA2 subtype (46% of total ANCA IgA) and presence of the secretory component concurrently. Presence of ANCA IgA was associated with disease-specific clinical characteristics (Child-Pugh stage and presence of ascites, p<0.001). During a 2-year follow-up period, risk of infections was higher among patients with ANCA IgA compared to those without (41.8% vs. 23.4%, p<0.001). ANCA IgA positivity was associated with a shorter time to the first infectious complication (pLogRank <0.001) in Kaplan-Meier analysis and was identified as an independent predictor in multivariate Cox-regression analysis (HR:1.74, 95% CI: 1.18-2.56, p=0.006). CONCLUSIONS: Presence of IgA type ANCA is common in cirrhosis. Involvement of gut mucosal immune system is in center of their formation and probably reflects sustained exposure to bacterial constituents.

Natural infection of baboons by Entamoeba histolytica elicits anti- gal-lectin heavy subunit IgA and IgG antibodies with shared epitope specificity to that of humans.

Non-human primates, such as baboons (Papio hamadryas anubis), are natural hosts for Entamoeba species; infections can be asymptomatic or result in invasive lethal disease. It was sought to determine whether following natural infection by Entamoeba. histolytica, baboon anti-amebic antibodies recognized native Gallectin, a recombinant portion of the lectin heavy subunit (designated LC3) and specific heavy subunit epitopes; we compared the specificity of anti-amebic antibodies from baboons to that of humans following asymptomatic E. histolytica infection or cure of amebic liver abscess (ALA). Female baboons (n=54), aged one to three years of age and living in captivity were screened for infection by real time PCR. E. histolytica infection was found in 37 baboons and was associated with serum anti-LC3 IgG (73%) and anti-LC3 IgA (46%) or intestinal anti-Gal-Lectin IgA antibody responses (49%), p<0.021 for each compared to that observed with baboons having an E. dispar infection (n=10) or uninfected baboons (n=7). The ELISA OD reading for anti-LC3 or anti-lectin antibodies correlated strongly with the presence of a PCR CT value indicative of E. histolytica infection. In humans with asymptomatic E. histolytica infection or those recently cured of ALA, 63% and 57% had serum anti- LC3 IgA and 65% and 57% had serum anti-LC3 IgG antibodies respectively. Epitope- specific synthetic peptides were used as capture antigens in ELISA; for baboons that possessed anti-LC3 and anti-lectin antibodies, 74% had anti-peptide IgG or IgA antibodies, compared to 86% of asymptomatic humans and 92% of ALA subjects(P>0.05).

Beneficial role of endogenous immunoglobulin subclasses and isotypes in septic shock.

PURPOSE: There is increasing evidence on the relationship between endogenously produced immunoglobulins and the clinical outcome in septic shock (SS). MATERIALS AND METHODS: Levels of immunoglobulin G (IgG) subclasses, immunoglobulin A (IgA), immunoglobulin M (IgM), and immunoglobulin E were measured in plasma from 42 patients with SS and in 36 patients with systemic inflammatory response syndrome at diagnosis. Association of immunoglobulins levels with disease severity and outcome was evaluated. RESULTS: Eighteen patients with SS finally died. Both patients with systemic inflammatory response syndrome and SS showed subnormal levels of total IgG, IgG2, and IgM. Patients with SS who died showed the lowest levels of total IgG and IgG1. Total IgG, IgG1, IgG2, IgG3, IgG4, and IgA correlated inversely with Acute Physiology and Chronic Health Evaluation II score in SS. Univariate Cox regression analysis showed that levels of IgG1, IgG2, IgG3, IgM, IgA, and total IgG were inversely associated to the probability of death at 28 days. Multivariate analysis showed that IgG1, total IgG, IgM, and IgA behaved as independent protective factors against mortality (hazard ratio, P): 0.23, 0.026; 0.16, 0.028; 0.11, 0.042; 0.05, 0.010, respectively, whereas IgG3 showed a protective trend also. CONCLUSIONS: Our study evidenced that, in addition to IgG1, other major endogenous immunoglobulins isotypes and subclasses seem to play a beneficial role in SS.

Profile of anti-Leishmania antibodies related to clinical picture in canine visceral leishmaniasis.

This research investigated the profile of anti-Leishmania antibodies in different clinical forms of canine visceral leishmaniasis (CVL). Naturally infected dogs were divided into two groups: subclinical dogs (SD, n=10) and clinical dogs (CD, n=68). Non-infected dogs (ND, n=7) comprised the negative control group. The humoral response was evaluated by the profile of total IgG, IgG1, IgG2, IgM, IgA and IgE, determined by ELISA. Infected animals showed increased levels of total IgG, IgA and IgE in addition to IgG1 and IgG2 in groups SD and CD, when compared with group ND. Furthermore, it was observed that IgG2 and IgM were correlated with symptomatology, while total IgG, IgG1 and IgA were negatively correlated and IgE showed no correlation. It follows that serum levels of IgG2 anti-Leishmania are correlated with typical clinical signs of disease. Furthermore the determination of specific anti-Leishmania antibodies could be an important tool in monitoring CVL clinical picture.

Human memory B cells originate from three distinct germinal center-dependent and -independent maturation pathways.

Multiple distinct memory B-cell subsets have been identified in humans, but it remains unclear how their phenotypic diversity corresponds to the type of responses from which they originate. Especially, the contribution of germinal center-independent responses in humans remains controversial. We defined 6 memory B-cell subsets based on their antigen-experienced phenotype and differential expression of CD27 and IgH isotypes. Molecular characterization of their replication history, Ig somatic hypermutation, and class-switch profiles demonstrated their origin from 3 different pathways. CD27⁻IgG⁺ and CD27⁺IgM⁺ B cells are derived from primary germinal center reactions, and CD27⁺IgA⁺ and CD27⁺IgG⁺ B cells are from consecutive germinal center responses (pathway 1). In contrast, natural effector and CD27⁻IgA⁺ memory B cells have limited proliferation and are also present in CD40L-deficient patients, reflecting a germinal center-independent origin. Natural effector cells at least in part originate from systemic responses in the splenic marginal zone (pathway 2). CD27⁻IgA⁺ cells share low replication history and dominant Igλ and IgA2 use with gut lamina propria IgA+ B cells, suggesting their common origin from local germinal center-independent responses (pathway 3). Our findings shed light on human germinal center-dependent and -independent B-cell memory formation and provide new opportunities to study these processes in immunologic diseases.

Monoclonal gammopathy associated with multiple myeloma and visceral leishmaniasis in the dog: a comparison of two cases.

The term monoclonal gammopathy (MG) suggests the presence of clonal immunoglobulins in blood serum that are recognized as narrow spikes in the beta and/or gamma region of the electrophoretic pattern of serum. In the dog, MG is rare and is associated with a heterogeneous group of diseases that include multiple myeloma (the most common source of MG) as well as infectious and chronic inflammatory diseases such as Leishmaniasis. In this paper, two cases of MG are described: the first case is associated with multiple myeloma of monoclonal component type IgA/lambda, with the latter rare in dogs, and the second case involves MG that developed 3 years after an initial diagnosis of Leishmaniasis.

[IgA subclass and IgA deficiency].

There are two subclasses of IgA, IgA1 and IgA2, and its heavy chains are encoded by two different genes, alpha1 and alpha2 genes. These two subclasses play important roles in the first line of defense, and the amount ratio of these molecules in secretions varies. IgA deficiency (IgAD) is the most common immunodeficiency, however the pathogenesis in most cases of IgAD is unknown. The class switch disorder in IgA producing B lymphocytes is one of the important factors in IgAD patients. The decreased expression levels of Ialpha germline transcripts before a class switch may be the cause of selective IgAD. The alpha1 and alpha2 gene expression levels are low in most IgAD patients. Using RT-PCR method in which alpha1 and alpha2 mRNAs can be separately evaluated, we identified the second case of alpha1 gene deletion in Japan. Longitudinal change in the serum IgA of the patient with alpha1 gene deletion showed the pattern of the partial IgAD. Patients with alpha1 gene deletion can be considered as having partial IgAD.

A T cell-dependent mechanism for the induction of human mucosal homing immunoglobulin A-secreting plasmablasts.

Mucosal immunoglobulin A (IgA) secreted by local plasma cells (PCs) is a critical component of mucosal immunity. Although IgA class switching can occur at mucosal sites, high-affinity PCs are optimally generated in germinal centers (GCs) in a T cell-dependent fashion. However, how CD4(+) helper T cells induce mucosal-homing IgA-PCs remains unclear. Here, we show that transforming growth factor beta1 (TGFbeta1) and interleukin 21 (IL-21), produced by follicular helper T cells (Tfh), synergized to generate abundant IgA-plasmablasts (PBs). In the presence of IL-21, TGFbeta1 promoted naive B cell proliferation and differentiation and overrode IL-21-induced IgG class switching in favor of IgA. Furthermore, TGFbeta1 and IL-21 downregulated CXCR5 while upregulating CCR10 on plasmablasts, enabling their exit from GCs and migration toward local mucosa. This was supported by the presence of CCR10(+)IgA(+)PBs in tonsil GCs. These findings show that Tfh contribute to mucosal IgA. Thus, mucosal vaccines should aim to induce robust Tfh responses.

Incomplete assembly of IgA2m(2) in Chinese hamster ovary cells.

Myeloma and Chinese hamster ovary (CHO) cells are frequently used for the production of recombinant antibodies. With increasing interest in producing recombinant IgA for protection against infectious agents, it is essential to characterize the IgA produced in these cells. Here we show that while myeloma cells secrete IgA2m(2) predominantly as H(2)L(2), CHO cells secrete H(2)L and H(2) in addition to fully assembled H(2)L(2). When the CHO cells also synthesize J chain and secretory component (SC), polymeric IgA and secretory IgA in which SC is disulfide bonded to the polymeric IgA are produced. Blocking cysteines on purified IgA2m(2) protein by alkylating with iodoacetamide stabilizes the disulfide bonds between the H and L chains suggesting that the disulfide bonds between H and L chains are unstable. Taken together our results suggest that the covalent assembly of IgA2m(2) is different in myeloma and CHO cells.