Characterization of five mouse monoclonal antibodies to apolipoprotein[a] from human Lp[a]: evidence for weak plasminogen reactivity

We describe the development of five murine monoclonal antibodies (14A12, 39A1, 53A9, 73A7, and 128A6) specific to human apolipoprotein[a] (Mr approximately 570,000), and their characterization by a number of procedures including cotitration, competition and inhibition enzyme-linked immunosorbent assays (ELISA), immunoblotting of native lipoproteins and of SDS-solubilized apolipoproteins electrophoresed in polyacrylamide gels, and dot immunobinding assays. The patterns of immunoreactivity of these antibodies were similar. Each reacted in ELISA assays and upon electroimmunoblotting with purified apo[a], with apo[a] liberated by reduction of Lp[a], and with delipidated Lp[a] solubilized in SDS, but by contrast, they reacted with native Lp[a] to a significant degree only upon electroimmunoblotting. No reactivity was seen with LDL-apoB-100 or with other apolipoproteins. The cross-reactivity of these antibodies with the homologous protein, plasminogen, was examined by comparison of the amount of plasminogen or apo[a] required for 50% inhibition of antibody binding to apo[a], and by an ELISA assay. The inhibition assay showed reactivity with plasminogen to be 37- to 50-fold lower than with apo[a], while dot immunobinding showed the lower limit of detection of plasminogen and of apo[a] to be approximately 320 and 31 micrograms, respectively. In an ELISA sandwich assay based on monoclonal antibodies LHLP-1, 14A12, and 53A9, the lower limit of Lp[a] detection (approximately 1 ng/ml protein) was about 100-fold less than that of plasminogen. Chemical modification of apo[a] revealed a significant contribution of arginine residues to the epitopes of 14A12, 39A1, and 53A9. Modification of cysteine residues with iodoacetamide was without effect, thereby distinguishing these antibodies from LHLP-1. Each antibody reacted with the six major size forms of apo[a] (Mr approximately 450,000-750,000) in immunoblots of human sera electrophoresed in SDS-polyacrylamide gels. Marked heterogeneity in apo[a] phenotype was detected and both single and double band phenotypes were observed in a randomized study. Cotitration and competition binding studies showed varying degrees of interaction between all five epitopes, with the exception of 128A6 which appeared to be independent of 39A1 and 53A9 (and vice versa). These data suggest that our five monoclonal antibodies recognize epitopes on apolipoprotein[a] that are exposed and accessible on the native Lp[a] particle. We conclude that our monoclonal antibodies recognize a specific region of apo[a], and that this region undergoes a conformational change upon adsorption of Lp[a] to plastic thereby diminishing epitope recognition.(ABSTRACT TRUNCATED AT 400 WORDS)     

Interaction of LDL, Lp[a], and reduced Lp[a] with monoclonal antibodies against apoB

Five monoclonal antibodies (2A, 9A, 6B, L3, L7) produced in mice against human apolipoprotein B were investigated by competitive and inhibitive electroimmunoassay (EIA) for their reactivity with low density lipoprotein (LDL), lipoprotein[a] (Lp[a]), and reduced Lp[a]. All of the antibodies reacted with apoB of the different lipoproteins indicated by very similar slopes of the binding curves. None of them gave a positive reaction with apolipoprotein[a]. The amount of apoB required for 50% inhibition of antibody binding varied for the different antibodies and lipoproteins. Antibody 9A showed almost the same affinity for LDL, Lp[a], and reduced Lp[a]. Antibodies 2A and 6B bound about twofold better to LDL and reduced Lp[a] than to untreated Lp[a]. Antibodies L3 and L7 needed nearly threefold higher amounts of Lp[a]-apoB for 50% inhibition of antibody binding than of apoB of LDL and reduced Lp[a]. The amount of apoB required for 50% inhibition of antibody binding was somewhat higher in inhibitive assay than in competitive assay. We suggest that apo[a] covers certain epitopes of apoB in native Lp[a] leading to a reduced reaction with the monoclonal antibodies. However, it could also be that the binding of the [a]antigen to apoB via disulfide bridges causes profound conformational changes of the apoB region exposed to the surface.     

A selective bi-site immunoenzymatic procedure for human Lp[a] lipoprotein quantification using monoclonal antibodies against apo[a] and apoB

A selective bi-site ELISA assay procedure for quantification of Lp[a] lipoprotein in human plasma based on linkage of apo[a] to apoB is described. The lipoproteins referred to as apo[a]:B were captured by a mixture of two anti-apo[a] monoclonal antibodies (K07, K09) and were revealed by a mixture of six anti-apoB monoclonal antibodies coupled to peroxidase. Since apo[a] and plasminogen have striking similarities in protein structure, the selective binding of Lp[a]:B in our assay depended upon the marked difference in affinity of the K07 and K09 mixture for Lp[a]:B (Kd = 0.32 x 10(-10) M) versus plasminogen (Kd = 0.47 x 10(-7)M). The high sensitivity (the Lp[a]:B working range 0.06-0.40 micrograms/ml) and the use of anti-apoB as antibody tracer added to the selectivity of the assay. The expression of K07 and K09 epitopes determined by competitive inhibition method and the reactivity of Lp[a]:B particles measured by bi-site ELISA were similar on individual lipoproteins, independent to their plasma levels. The assay is precise, and intra- and interassay coefficients of variation were 4.7% and 9.6%, respectively. It yields quantitative Lp[a]:B values that correlate highly with Lp[a] levels obtained by electroimmunoassay with polyclonal antibody (r = 0.73) or with Lp[a] levels measured by the other bi-site ELISA using only K07 and K09 antibodies (r = 0.96). However, upon analyzing each individual plasma with an arbitrary Lp[a]-cut off of 15 mg/dl, evidence of the qualitative aspect of the lipoprotein was obtained. The group with Lp[a] less than 15 mg/dl had higher frequency of subjects (65%) with the ratio Lp[a]/Lp[a]:B above 1.5.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vitro generated human monoclonal trinitrophenyl-specific B cell lines. Evidence that human and murine anti-trinitrophenyl monoclonal antibodies cross-react with Escherichia coli beta-galactosidase

Stable human antigen-specific monoclonal B cell lines were established without prior in vivo immunization. This was accomplished by expanding the anti-trinitrophenyl (TNP) B cells in vitro with the antigen TNP-Brucella abortus and then immortalizing them with Epstein-Barr virus. Five anti-TNP clones were selected by sequential limiting dilution. All five anti-TNP clones secreted IgM kappa antibodies. When tested against a panel of self and environmental antigens, all five anti-TNP clones exhibited cross-reactivity with an Escherichia coli-derived beta-galactosidase. To determine whether this was a more general phenomenon, a panel of murine monoclonals were tested and found to bind to beta-galactosidase. It is therefore possible that human and murine anti-TNP beta cell responses reflect reactivity against an environmental antigen, namely an epitope present on E. coli-derived beta-galactosidase. This approach of expanding human antigen-specific B cells by antigen stimulation in vitro, with a T-independent hapten-carrier conjugate before Epstein-Barr virus transformation, may prove useful in the development of human monoclonals for therapeutic purposes.     

Heterogeneity of human lipoprotein Lp[a]: cytochemical and biochemical studies on the interaction of two Lp[a] species with the LDL receptor

Human Lp[a] can be fractionated into two species with different affinities for lysine-Sepharose. Forty to 81% of the total Lp[a] in the density fraction 1.055-1.15 g/ml from five individuals was retained by this affinity column. The remaining unretained Lp[a] species with no apparently functional lysine binding site was similar to the retained species in its electrophoretic mobility, lipid, protein, and apolipoprotein composition, and the heterogeneity was not related to apo[a] size polymorphism. Interaction of the two species with the low density lipoprotein (LDL) receptor was studied in human fibroblasts. Using gold-labeled lipoproteins and an immunochemical procedure, both Lp[a] species could be located in clusters on the cell surface, but the extent of labeling was far lower than that seen with LDL. Both Lp[a] variants were less effective than LDL in 1) down-regulation of LDL-receptor activity; 2) suppression of cellular sterol synthesis; and 3) stimulation of cholesteryl ester formation in human fibroblasts. Although degradation of both species of Lp[a] by the perfused rat liver was stimulated after estrogen induction of hepatic LDL-receptor activity, the stimulation amounted to only a quarter of that seen with LDL. The heterogeneity of Lp[a] with respect to the ability to bind epsilon-aminocarboxylic acid will need to be considered in studying the physiological role of this lipoprotein. Both Lp[a] species exhibited a similar interaction with the LDL-receptor in vitro, and confirmed previous investigations that Lp[a] is only a poor ligand for the LDL-receptor.     

Preparation and characterization of monoclonal antibodies to human apolipoprotein E]

Monoclonal antibodies to human apolipoprotein E (apoE) were prepared and characterized. Antibodies of 3D12F11 clone were shown to be specific to apoE and belong to IgG1 subclass. Dissociation constant of antigen-antibody complex in solution was determined as 3.5 +/- 0.5 nM. The antibodies interacted neither heparin- nor lipid-binding sites of human apoE molecule. The obtained antibodies may be useful for metabolic and structural investigations of apoE as well as for clinical studies.     

Toxoplasma gondii: characterization of monoclonal antibodies that recognize rhoptries

We have previously reported on a series of monoclonal antibodies that recognize the rhoptries of Toxoplasma gondii and that interfere with the action of penetration enhancing factor. The antibodies immunoprecipitate several related antigens from [35S]methionine-labeled parasites that range in size from 60 to 43 kDa. By immunoblot, one of the antibodies reacts with the 60 kDa protein in the presence of protease inhibitors. Trypsin digestion of the antigen destroyed antigenic reactivity indicating that the 60 kDa antigen is a protein. The antigen was stable to periodate oxidation and failed to react with Schiff's reagent, indicating that the antigen contains little or no carbohydrate. Two-dimensional gel electrophoresis followed by immunoblot showed that the antigen recognized by Tg 49 was an acidic protein with an approximate pI of 5.8.     

Three monoclonal antibodies against measles virus F protein cross-react with cellular stress proteins.

In a group of 11 monoclonal antibodies specifically reacting with the measles virus fusion protein, three antibodies also immunoprecipitated other proteins, in particular a 79,000-molecular-weight protein from virus-infected cells. The cross-reacting 79,000-molecular-weight protein was shown to be a virus-induced host stress protein. This protein could be induced by (i) different paramyxoviruses, (ii) heat shock of uninfected HeLa cells, and (iii) 2-deoxyglucose, tunicamycin, or L-canavanine treatment of different mammalian cell lines. Immunofluorescence of stressed HeLa cells localized the cross-reacting host protein(s) mainly in the cytoplasm. The significance of these results in relation to autoimmunity is discussed.     

Characterization of monoclonal antibodies to Acanthamoeba myosin-I that cross-react with both myosin-II and low molecular mass nuclear proteins

We characterized nine monoclonal antibodies that bind to the heavy chain of Acanthamoeba myosin-IA. Eight of these antibodies bind to myosin-IB and eight cross-react with Acanthamoeba myosin-II. All but one of the antibodies bind to a 30-kD chymotryptic peptide of myosin-IA that derives from the COOH terminus of the molecule, and to tryptic peptides as small as 17 kD, hence these epitopes are clustered closely together on the heavy chain. None of the antibodies prevent heavy chain phosphorylation by myosin-I heavy chain kinase. One antibody inhibits the K+-EDTA ATPase activity and three antibodies inhibit the actin- activated Mg++-ATPase activity of myosin-I under the set of conditions that we tested. When fluorescent antibody staining of both whole cells and isolated nuclei is done, several of these monoclonal antibodies react strongly with nuclei. These antibodies also stain the cytoplasmic matrix, especially the cortex near the plasma membrane. All nine of the monoclonal antibodies bind to polypeptides of 30-34 kD that are highly enriched in nuclei isolated from Acanthamoeba. There is no myosin-I in the isolated nuclei, so the 30-34-kD polypeptides, not myosin-I, are responsible for the nuclear staining.     

Isolation of human monoclonal antibodies that neutralize human rotavirus

A human antibody library constructed by utilizing a phage display system was used for the isolation of human antibodies with neutralizing activity specific for human rotavirus. In the library, the Fab form of an antibody fused to truncated cp3 is expressed on the phage surface. Purified virions of strain KU (G1 serotype and P[8] genotype) were used as antigen. Twelve different clones were isolated. Based on their amino acid sequences, they were classified into three groups. Three representative clones-1-2H, 2-3E, and 2-11G-were characterized. Enzyme-linked immunosorbent assay with virus-like particles (VLP-VP2/6 and VLP-VP2/6/7) and recombinant VP4 protein produced from baculovirus recombinants indicated that 1-2H and 2-3E bind to VP4 and that 2-11G binds to VP7. The neutralization epitope recognized by each of the three human antibodies might be human specific, since all of the antigenic mutants resistant to mouse monoclonal neutralizing antibodies previously prepared were neutralized by the human antibodies obtained here. After conversion from the Fab form of an antibody into immunoglobulin G1, the neutralizing activities of these three clones toward various human rotavirus strains were examined. The 1-2H antibody exhibited neutralizing activity toward human rotaviruses with either the P[4] or P[8] genotype. Similarly, the 2-3E antibody showed cross-reactivity against HRVs with the P[6], as well as the P[8] genotype. In contrast, the 2-11G antibody neutralized only human rotaviruses with the G1 serotype. The concentration of antibodies required for 50% neutralization ranged from 0.8 to 20 micro g/ml.     

Biological characterization of human monoclonal antibodies to rabies virus.

Rabies virus antigen-specific human monoclonal antibodies (MAbs) that recognized either viral glycoprotein, ribonucleoprotein, or matrix proteins were generated. Only glycoprotein-specific MAb neutralized a variety of rabies viruses and protected laboratory rodents against lethal rabies virus infection. The determinant recognized by this MAb does not appear to reside in previously defined antigenic sites of the viral glycoprotein.     

Isolation and characterization of human monoclonal antibodies to digoxin.

Fab preparations of sheep polyclonal anti-digoxin Abs have proven useful for reversal of the toxic effects of digoxin overdoses in patients. Unfortunately, the use of foreign species proteins in humans is limited because of the potential for immunological responses that include hypersensitivity reactions and acute anaphylaxis. Immunization of recently developed transgenic mice, whose endogenous micro heavy and kappa light chain Ig genes are inactivated and which carry human Ig gene segments, with a digoxin-protein conjugate has enabled us to generate and isolate eight hybridoma cell lines secreting human sequence anti-digoxin mAbs. Six of the mAbs have been partially characterized and shown to have high specificity and low nanomolar affinities for digoxin. In addition, detailed competition binding studies performed with three of these mAbs have shown them to have distinct differences in their digoxin binding, and that all three structural moieties of the drug, the primary digitoxose sugar, steroid, and five-member unsaturated lactone ring, contribute to Ab recognition.     

Production and characterization of monoclonal antibodies that specifically bind to phosphatidylcholine.

A series of monoclonal antibodies (mAbs) that react with phosphatidylcholine (PC) were established. All mAbs were highly specific to PC and no cross-reaction with other phospholipids were observed. The results obtained with two typical monoclonal antibodies, JE-1 and JE-8, were described. The analysis using synthetic PC analogs with modified polar head groups showed that the methyl groups on the quaternary nitrogen of the choline moiety were important for the binding. Each mAbs showed distinct acyl chain specificities of the PC molecules, and JE-1 showed considerable reactivity with PC with saturated fatty acids, whereas JE-8 could not react with the PC. Both mAbs bound to PC with unsaturated fatty acids, but showed distinct reactivity profiles. Both mAbs reacted only weakly with water-soluble haptens such as phosphorylcholine and L-alpha-glycerophosphocholine, suggesting that the hydrophobic moiety of the PC molecule is important for the maximum affinity. The interaction between the mAbs and the hydrophobic moieties of PC molecules was further studied by analyzing the effect of the mAbs on the activities of phospholipase A2 and phospholipase C. JE-1 inhibited both enzyme activities, while JE-8 inhibited only the phospholipase C activity, indicating that JE-1 interacts more thoroughly with the hydrophobic region of the PC molecule than JE-8 does.     

Immunopurification and characterization of human alpha-L-iduronidase with the use of monoclonal antibodies.

alpha-L-Iduronidase from human liver was purified by a three-step five-column procedure and by immunoaffinity chromatography with a monoclonal antibody raised against purified enzyme. Seven bands identified by staining with Coomassie Blue had molecular masses of 74, 65, 60, 49, 44, 18 and 13 kDa and were present in both preparations of the liver enzyme. However, relative to the immunopurification procedure, alpha-L-iduronidase purified by the five-column procedure was considerably enriched in the 65 kDa polypeptide band. The seven bands were identified by Western-blot analysis with two different monoclonal antibodies raised against alpha-L-iduronidase. The chromatographic behaviour of alpha-L-iduronidase on the antibody column was dependent upon the quantity of enzyme loaded. Above a particular load concentration a single peak of enzyme activity was eluted, whereas at load concentrations below the critical value alpha-L-iduronidase was eluted in two peaks of activity, designated form I (eluted first) and form II (eluted second). The following properties of the two forms of alpha-L-iduronidase were determined. (1) The two forms from liver were composed of different proportions of the same seven polypeptides. (2) When individually rechromatographed on the antibody column, each form from liver shifted to a more retarded elution position but essentially retained its chromatographic behaviour relative to the other form. (3) Forms I and II of liver alpha-L-iduronidase showed no difference in their activities towards disaccharide substrates derived from two glycosaminoglycan sources, heparan sulphate and dermatan sulphate. (4) The native molecular size of forms I and II of liver alpha-L-iduronidase was 65 kDa as determined by gel-permeation chromatography. (5) Immunoaffinity chromatography of extracts of human lung and kidney resulted in the separation of alpha-L-iduronidase into two forms, each with different proportions of the seven common polypeptide species. (6) Lung forms I and II were taken up readily into cultured skin fibroblasts taken from a patient with alpha-L-iduronidase deficiency. Liver forms I and II were not taken up to any significant extent. Lung form II gave intracellular contents of alpha-L-iduronidase that were more than double those of normal control fibroblasts, whereas lung form I gave contents approximately equal to normal control values. We propose that all seven polypeptides are derived from a single alpha-L-iduronidase gene product, and that different proportions of these polypeptides can function as a single alpha-L-iduronidase entity.(ABSTRACT TRUNCATED AT 400 WORDS)     

Interspecies cross-reactivity of monoclonal antibodies to various epitopes of human plasminogen

The immunological cross-reactivities of three conformationally specific monoclonal antibodies to distinct epitopes on human plasminogen toward plasminogens purified from 14 additional species have been examined. Antibody 10-F-1, which is produced against an epitope on the kringle 4 region of human plasminogen, shows a high degree (greater than 80%) of cross-reactivity against baboon, goat, monkey, ovine, and rabbit plasminogens; more limited (20-50%) cross-reactivity against bovine, equine, goose, guinea pig, mouse, rat, and porcine plasminogens; and little comparable cross-reactivity against canine and chicken plasminogens. Antibody 10-H-2, generated to an epitope of the kringles 1-3 region of human plasminogen, shows extensive cross-reactivity (72%) only toward monkey plasminogen, more limited (22-35%) cross-reactivity toward equine and rabbit plasminogens, and much less cross-reactivity toward any other of the above plasminogens. Antibody 10-V-1, also produced against an epitope on the kringle 1-3 region of human plasminogen, which is distinct from the 10-H-2 epitope, shows extensive cross-reactivity (72-100%) with baboon, monkey, and rabbit plasminogens; more limited cross-reactivity with equine (48%) and mouse (28%) plasminogens; and a low level of such reactivity with the remaining plasminogens. These studies show that the extent of interspecies cross-reactivity of various plasminogens greatly depends upon the epitope in question. The K4 region of these molecules appears more extensively conserved than the K1-3 region, at least in regard to the particular epitopes examined in this study.     

The development of porcine zona pellucida using monoclonal antibodies: I. Immunochemistry and light microscopy

We established three monoclonal antibodies (Mabs) against the zonae pellucidae (ZP) of porcine oocytes, named STA-1, STA-2, and STA-3, and eventually we determined that they all reacted with the isolated ZP. Based on Western blotting without 2-mercaptoethanol (2-ME), STA-1 reacted with the 80,000-110,000 Mr component, STA-2 with the 42,000-63,000 Mr component, and STA-3 with the 40,000-80,000 Mr component of ZP. We immunohistochemically specified the components of porcine ZP reactive with the three Mabs during the course of follicular development. Each Mab reacted with both the ZP and the interfollicular cell space (IFCS). One ZP component, reactive with STA-2 and STA-3, was first produced in the primordial follicle and was not found at the cumulus follicle stage, which corresponds to the stage of large antral follicles more than 5 mm in diameter. Another ZP component, reactive with STA-1, was not produced until the secondary follicle stage, and was never found at the antral follicle stage. These results suggest that each ZP component is produced and secreted at a specific stage or stages of folliculogenesis.     

Analysis of human C8 with monoclonal antibodies. Characterization of a monoclonal antibody that recognizes free C8 alpha-gamma subunit

The eighth component of human C is essential for the formation of the membranolytic C attack complex. C8 has a unique structure in that two covalently linked chains, C8 alpha and C8 gamma, are associated non-covalently with the third chain, C8 beta. In order to study the structure and assembly of the C8 molecule, a panel of mAb has been produced against the C component C8. Eight of these mAb had reactivity to the C8 alpha-gamma subunit, whereas four reacted with C8 beta. One of the C8 alpha-gamma mAb, C8A2, had specificity for an epitope on the C8 alpha-chain and exhibited no cross-reactivity to any of the other terminal C components, including C8 beta. C8A2 inhibited the hemolytic activity of the C8 alpha-gamma subunit but had no effect on the activity of fluid phase whole C8 or C8 within membrane-bound C5b-8. Functional experiments suggest that C8A2 inhibits C8 alpha-gamma activity by interfering with its interaction with the C8 beta-chain. In an enzyme immunoassay using the C8A2 mAb, free C8 alpha-gamma subunit could be detected in both homozygous and heterozygous C8 beta-deficient serum. However, only low level binding was observed when homozygous C5- and C7-deficient sera were tested. Thus the mAb, C8A2, recognizes an epitope expressed on the C8 alpha-gamma subunit but not on intact C8 and can detect free C8 alpha-gamma in the presence of native C8.     

Immunochemical characterization of human liver and heart ferritins with monoclonal antibodies

A library of 27 murine monoclonal antibodies was obtained by using human liver and heart ferritins as immunogens. The specificity of the antibodies for the two ferritins and their subunits was studied with five different methods. The antibodies elicited by the liver ferritin bound preferentially the immunogen and were specific for the L subunit. Some antibodies elicited by the heart ferritin had characteristics similar to the anti-liver antibodies, other ones bound preferentially the heart over the liver ferritin and were specific for the H subunit. Only two antibodies were able to bind both ferritins and subunits. Some anti-H and anti-L chain antibodies were used to develop and compare four types of immunoassay to quantitate isoferritins. The results indicate that heart ferritin is immunologically more heterogeneous than liver, the H and L subunits having large immunological differences with few, if any, identical epitopes; and that that the architecture of the immunoassays have a strong influence on the crossreactivity of the antibodies with the two isoferritins, probably because H and L chains are not arranged randomly in the assembled protein.     

Structural characterization of human melanoma-associated antigen p97 with monoclonal antibodies

Monoclonal antibodies were used to study p97, a human melanoma-associated antigen (MAA). Four hybridomas, designated 4.1, 96.5, 118.1, and 8.2, were obtained by fusing mouse myeloma cells with spleen cells from mice immunized with human melanoma cells. Antibodies 4.1 and 8.2 were IgG1; antibodies 96.5 and 118.1 were IgG2a. Sequential immunoprecipitation (IP) and sodium-dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that all 4 antibodies recognized the same 97 kilodalton (kD) protein. Binding studies with 125I-labeled antibody showed that antibodies 4.1 and 96.5 bound the same epitope, p97a. Antibodies 118.1 and 8.2 defined epitopes p97b and p97c, respectively. Six monoclonal antibodies (M17, L1, L10, R10, I12, and K5) specific for gp95, a kD melanoma cell surface glycoprotein were also tested. Sequential IP showed that these antibodies bound p97; p97 and gp95 are thus identical. Binding studies showed that antibody m17 bound epitope p971, and antibodies L1, L10, and R19 bound epitope p97c. Antibodies I12 and K5 defined 2 other epitopes, p97d and p97e, respectively. SDS-PAGE under nonreducing conditions indicated that p97 is monomeric, probably with intrachain disulfide bonds. Cell-surface labeling of sialic acid residues and neuraminidase digestion showed that p97 is a sialoglycoprotein. Digestion of p97 with papain or trypsin produced a stable 40 kD fragment, which expressed epitopes p971, p97b, and p97c, but not p97d or p97e.     

Development of monoclonal antibodies that recognize antigens associated with human cervical carcinoma

Six monoclonal antibodies, generated by immunization of mice with human cervical carcinoma cells maintained in tissue culture or with cells from fresh tumor tissue, reacted specifically with the malignant cells in 71% to 90% of the tumor tissue imprints and cervical smears containing neoplastic cells but not with normal cervical epithelial cells in smears from 21 to 23 healthy donors. Antibody CE 402 bound to epithelial cells associated with regeneration in 2 of the 23 normal smears tested. Considerable heterogeneity of antibody binding by malignant cells was observed. Antibody CE 400 was the most reactive, binding to more than 50% of the tumor cells in all reactive specimens. Five of these monoclonal antibodies detected protein antigens in the 80 K to 110 K molecular weight range. Our studies demonstrate the feasibility of producing monoclonal antibodies with selected specificity for cervical carcinoma. These antibodies may be of considerable diagnostic value.