Biomedical subjects

P Perlmann

Publications and source records attributed to P Perlmann.

At least 109 records · Page 6Linked to original sources

PubMed1989

Studies on the role of CD43 in human B-cell activation and differentiation.

The monoclonal antibody (MoAb) B1B6 to human leucocyte sialoglycoprotein, CD43, induces aggregation of T cells and delivers progression signals early during activation of both T and B cells in the presence of primary activators of protein kinase C. In this report we further studied the role of CD43 in human B-cell activation and differentiation. About 5-10% of resting tonsillar B cells are CD43+. In the presence of TPA or antibodies to CDw40, the proportions of CD43+ cells drastically increased. The expression was optimal on day 3 of culture, when up to 80% and 50%, respectively, were CD43+. Whereas MoAb B1B6 together with TPA induced a three- to fivefold higher proliferative response as compared to TPA alone, antibodies to CDw40 did not synergize with MoAb B1B6 in B-cell proliferation. Tonsillar populations depleted of CD43+ B cells responded with lower proliferation to TPA alone or to TPA and B1B6 or anti-CDw40 antibodies. MoAb B1B6 did not affect the production of IgM or IgG as induced by pokeweed mitogen in the presence of autologous T cells, from either peripheral blood or tonsillar B cells. Neither did it affect the IgG production from the CD43+ BSF-2 sensitive Epstein-Barr virus-transformed lymphoblastoid cell line CESS. The results show that CD43 is upregulated on B cells during activation. Furthermore, CD43+ B cells are included in the population which responds to signals delivered by TPA, anti-CD43 or anti-CDw40 antibodies, and the proliferation of this population is not merely due to an expansion of the small population of CD43+ cells present among these cells. Moreover, the epitopes recognized by MoAb B1B6 are not involved in the differentiation of and ultimate Ig-secretion from activated B cells.

Antibodies, Monoclonal↗

PubMed1989

Enhancement of human B-cell proliferation by a monoclonal antibody to CD43.

Monoclonal antibodies (MoAb) to human leucocyte sialoglycoprotein, CD43, have been shown to deliver mitogenic signals to human T cells or to enhance T-cell proliferation induced by concanavalin A, anti-CD3 antibodies or phorbol ester. In this paper, we studied the effects of anti-CD43 MoAb B1B6 on the activation of human B cells. Anti-CD43 MoAb B1B6 was not mitogenic by itself for human B cells. However, when added together with TPA, both resting and in vivo activated tonsillar B cells, containing 5-10% and about 35% CD43+ respectively, responded with three- to fivefold higher proliferation compared to that obtained with TPA alone. A peak in the proliferative response was reached on day 3. Optimal proliferation was obtained when the antibody was present from the start of culturing. Addition of MoAb B1B6 together with a calcium ionophore, ionomycin, did not induce B-cell proliferation. Neither did mAb B1B6 sustain the growth of B cells that were already in the cell cycle, i.e. precultured with phorbol ester (PDB) and ionomycin for 3 days. The results are similar to those obtained with antibodies to CD22 and CD23 and show that early progression signals are delivered to resting B cells through CD43 in the presence of primary activators of protein kinase C.

Adjuvants, Immunologic↗

PubMed1989

Persistent superphosphorylation of leukosialin (CD43) in activated T cells and in tumour cell lines.

CD43 (leukosialin) is a highly sialylated, single-chain molecule expressed on most human leucocytes. Regulatory signals appear to be transduced through the molecule as suggested by the ability of anti-CD43 antibodies to induce aggregation and proliferation of T cells and to enhance B-cell proliferation and natural killer cell activity. Activation of protein kinases is an essential event in signal transduction. We were therefore interested to study whether CD43 may function as a substrate for protein kinases during mitogenic activation of lymphocytes. We show that CD43 was rapidly superphosphorylated (within minutes) on serine residues following addition of phorbol ester (PMA) to peripheral blood lymphocytes. PMA treatment of the cells was not followed by rapid down-regulation of CD43. Activation of the lymphocytes by concanavalin A or anti-CD3 antibodies (OKT-3) also resulted in superphosphorylation of CD43. However, the phosphorylation was delayed as compared to that induced by PMA and was detected 3-4 h after the addition of the reagents. A plateau was reached after 24-48 h of stimulation. Interestingly, the high level of phosphorylation of CD43 was maintained in long-term cultures of T cells activated by various means. Furthermore, CD43 was found to be constitutively superphosphorylated (on serine and tyrosine) in continuously growing cell lines of T, B, and non-lymphoid origin. Taken together, the results suggest that CD43 has an important role during both early and late phases of T-cell activation and that modulation of its biochemical properties by protein kinases may be associated with progression through the cell cycle and with cellular growth.

Antigens, CD↗

PubMed1989

Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components.

The development of vaccines is presently receiving major attention in malaria research. As it is not possible to base malaria vaccines on the use of killed or attenuated organisms, the vaccines which are being developed are subunit vaccines in which the immunogens consist of defined parasite antigens or antigenic fragments. Since protective immunity to malaria involves both antibody-dependent and antibody-independent mechanisms, the immunogens in a subunit vaccine must have the capacity to induce relevant B- and T-cell responses in the majority of vaccinees. In turn, this requires good knowledge of these responses in humans who have acquired immunity through natural infection. In this paper we have summarized our recent work on the dissection into epitope-specific components of the human antibody response to the Plasmodium falciparum antigen Pf155/RESA, a recognized candidate for a vaccine against the asexual blood stages of this parasite. Epitope mapping of the antigen by means of short synthetic peptides led to the identification in several molecular regions of short amino acid sequences constituting linear and probably immunodominant B-cell epitopes. The antigenically most active region was located in the C-terminus of the molecule. This region, which consists of approximately 40 related, 4- or 8-amino acid long repeats, induced higher antibody concentrations in a larger number of malaria-immune donors than any of the other regions. A large fraction of these antibodies bound to short synthetic peptides representing the major repeat motifs of Pf155/RESA. Although these repeats are made up of closely related amino acid sequences, the antibody response to them was highly polyclonal, indicating the presence of several linear and probably also conformational epitopes which gave rise to a variety of cross-reacting as well as monospecific antibodies. Further analysis revealed that the levels of antibodies differing in specificity and/or avidity for different peptides varied independently of each other in individual donors. In an area (Liberia) where malaria transmission is holoendemic and perennial, these antibody profiles remained constant when individual donors were followed over several years. Since the C-terminal repeat region of Pf155/RESA is conserved in different P. falciparum strains, the results reflect differences in the genetic regulation of epitope-specific host responses rather than antigenic differences between infecting parasites. In donors living in an area with high but seasonal malaria transmission, antibody levels usually drop to lower levels when there is no transmission.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗

PubMed1989

Movement of a falciparum malaria protein through the erythrocyte cytoplasm to the erythrocyte membrane is associated with lysis of the erythrocyte and release of gametes.

Erythrocytes containing mature gametocytes of Plasmodium falciparum circulate in the blood until they are ingested by a mosquito, an event that triggers gametogenesis and lysis of the infected erythrocyte. It was previously shown that a parasite protein (Pf155/RESA) accumulates in the erythrocyte cytoplasm next to the parasitophorous vacuolar membrane (S. Uni, A. Masuda, M. J. Stewart, R. Nussenzweig, and M. Aikawa, Am. J. Trop. Med. Hyg., 36:481-488, 1987). Using a monoclonal antibody to Pf155/RESA and rabbit sera to two different repeat peptides of Pf155/RESA, we have studied the location of Pf155/RESA after induction of gametogenesis. Five minutes after triggering gametogenesis, the parasitophorous membrane no longer surrounded the parasite, bringing the parasite membrane in contact with the erythrocyte cytoplasm. Clear spaces appeared throughout the hemoglobin-rich host cytoplasm; Pf155/RESA was now localized in the cytoplasm directly surrounding the spaces. No membrane existed between the spaces and the erythrocyte cytoplasm. The spaces with surrounding Pf155/RESA protein extended to the erythrocyte membrane. After lysis of the erythrocyte membrane (15 min after triggering gametogenesis), the protein was distributed along the erythrocyte membrane and throughout the space between the gamete and the erythrocyte membrane. The mechanism by which Pf155/RESA remained aggregated around the spaces and its role in erythrocyte lysis are unknown. It is of interest that the parasite appeared to use the same molecule during invasion of erythrocytes and during release of gametes from infected erythrocytes.

Animals↗

PubMed1989

An epidemiological study of humoral and cell-mediated immune response to the Plasmodium falciparum antigen PF155/RESA in adult Liberians.

We investigated the seroreactivity and T cell reactivity against the Plasmodium falciparum antigen Pf155/RESA, different oligopeptides from the 3' and 5' repeat regions of the Pf155/RESA antigen, and crude Plasmodium falciparum antigens in 164 adult Liberians. We compared 2 long-term residential groups with high and low exposure to malaria. The seropositive rate to the peptides was significantly higher with increased exposure. There was no significant difference in response rates to the Pf155/RESA. This may indicate the level of persistent T cell memory in previously primed donors. The seropositive rates to 3 Pf155/RESA peptides and the rates measured by either 3H-thymidine incorporation or IFN-gamma release after stimulation with Pf155/RESA and the peptides were all lower in parasite positive individuals. Even low grade, asymptomatic parasitemia can impair the T cell response in vitro. The lower antibody response in parasite positive subjects may be explained by either antibody consumption or lower protection against malaria parasitemia in subjects with low concentrations of antibodies against the Pf155/RESA antigen.

Adult↗

PubMed1989

Phenotypic characterization of mononuclear white cells using finger-prick blood and light microscopy.

A technique for the enumeration of T cell subsets in capillary blood for use in the field is described. Mononuclear white cells were separated from 0.4 ml capillary blood, kept covered by culture medium on a Multitest slide, and incubated with monoclonal antibodies to the CD2, CD4, CD8, and interleukin-2 receptor antigens. Secondary and tertiary alkaline phosphatase conjugates were used for labeling. The substrate gave a distinct blue surface staining to the positive lymphocytes. Comparison was made with a conventional immunofluorescent antibody technique (r = 0.95).

Adult↗

PubMed1989

Surface antigens on equine sarcoid cells and normal dermal fibroblasts as assessed by xenogeneic antisera.

To characterise the expression of surface antigens on equine sarcoid cells compared to normal equine fibroblasts, immune sera were produced in rabbits against transformed cells of a virus-containing sarcoid cell line (Mc-1) and normal dermal fibroblasts, respectively. The specificities of the sera were analysed by antibody-dependent cellular cytotoxicity against 51Cr-labelled target cells using human lymphocytes as effector cells. Anti-Mc-1 antiserum induced strong cytotoxicity against transformed cells of two sarcoid cell lines (Mc-1 and Bay Mc-1), whereas the cytotoxicity against transformed cells of equine testis, or against cells grown in short term or primary cultures derived either from sarcoids or normal dermis was low or absent. In contrast, anti-normal dermal fibroblasts antiserum did not induce antibody-dependent cellular cytotoxicity against Mc-1 or Bay Mc-1 but reacted strongly against both normal fibroblasts and primary cultured cells derived from sarcoids. The apparent specificity of the anti-Mc-1-induced cytotoxicity was confirmed by absorption of the anti-serum with either normal dermal fibroblasts and equine testis cells or Mc-1 cells. Immunoprecipitation of 125I-surface labelled Mc-1 cells by absorbed anti-serum and analysis by SDS-PAGE and autoradiography revealed that anti-Mc-1 antibodies reacted with at least four surface components with a wide range of molecular weights. Immunofluorescence with rabbit anti-human beta 2mu serum indicated that Mc-1 cells in contrast to equine lymphocytes and fibroblasts did not express class 1 major histocompatibility complex antigens.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1988

Induction of aggregation and enhancement of proliferation and IL-2 secretion in human T cells by antibodies to CD43.

CD43 (large sialoglycoprotein) is a heavily glycosylated protein expressed on virtually all thymus-derived lymphocytes, on a subpopulation of B cells and on granulocytes. Recently, an anti-CD43 mAb (L10) was shown to induce proliferation in T cells comparable to that induced by anti-CD3. The L10 antibody was reported to react with both sialylated and desialylated CD43. In order to further elucidate the role of CD43 in various T cell functions we have studied the biologic properties of two other mAb (B1B6 and E11B, IgG1) directed against sialic acid-dependent epitopes on CD43. Addition of low amounts of antibody (5 to 10 ng/ml) to freshly isolated T cells or to T cell lines resulted in a rapid clustering of the cells. Fab fragments were also active albeit at a 10-fold higher concentration. Aggregation was dependent on active cell metabolism (inhibited by azide and at low temperatures), on the presence of divalent cations (Mg2+) and was inhibited by antibodies to CD18 but not by antibodies to CD11a (leukocyte function-associated Ag-1 alpha). B1B6 and E11B were poorly mitogenic when added alone in soluble form to PBL or to T cells. However, supernatants from cultures of PBL treated with B1B6 for 2 days contained IL-2 activity. No increase in the number of CD25+ cells was seen during the same period. Exogenously added IL-2 did not synergize with B1B6 or E11B in activation of PBL, whereas proliferation was significantly increased by the addition of the antibodies to activation systems with low endogenous production of IL-2 (PMA or soluble anti-CD3). The anti-CD43 antibodies amplified T cell proliferative responses induced by Con A or leukoagglutinin from Phaseolus vulgaris. F(ab')2 fragments enhanced proliferation significantly better than Fab fragments suggesting that cross-linking of CD43 molecules was an essential features of the amplifying signal. Compared with cultures activated by Con A alone, an increased number of CD25+ cells and of blast cells as well as an increased IL-2 production was observed in cultures activated by B1B6-Con A. The results indicate that regulatory signals, which may function to modify homo- or heterotypic T cell adhesion as well as autocrine production of IL-2, can be transduced through CD43.

Antibodies, Monoclonal↗

PubMed1988

MHC nonrestricted cytotoxic T cell clones with selective specificity from patients with transitional cell carcinoma (TCC) of the urinary bladder.

Lymphocytes from patients with transitional cell carcinoma (TCC) of the urinary bladder are more cytotoxic to bladder tumor cells than to a variety of control cells. This disease-related cytotoxicity has previously been shown to involve several mechanisms and different types of effector cells. To analyze further the nature of the effector cells operative in this system, peripheral blood lymphocytes from eight TCC patients were stimulated in vitro with TCC extract and cultured in the presence of interleukin 2 and allogeneic feeder cells. When tested for cytotoxicity in vitro on a target cell panel including both adherent and nonadherent cell lines, the lymphocytes killed a broad spectrum of targets in a major histocompatibility complex (MHC)-unrestricted fashion. When cloned by limiting dilution, clones were obtained which displayed a more restricted pattern of target cell killing. Some of the clones were highly but not exclusively selective for TCC-derived target cells. Phenotypically, these cells resembled mature T cells of CTL-type (CD8+/CD4-). They also expressed the CD3/5 T cell antigen receptor complex but target cell killing was not MHC-restricted. The results of various inhibition experiments suggested that the CD3/TCR complex was involved in the cytotoxicity exhibited by these effector cells. However, its precise role in target cell recognition and the identification of the tumor cell structures recognised by the effector cells require further studies.

Antigens, Differentiation, T-Lymphocyte↗

PubMed1988

Cloning of genomic fragment from Plasmodium chabaudi expressing a 105 kilodalton antigen epitope.

The 105 kDa antigen of Plasmodium chabaudi has many characteristics of the P. falciparum Pf 155 (RESA) molecule. A clone (pPC105e) from a P. chabaudi genomic library was isolated using immune screening with a 105 kDa antigen specific monoclonal antibody (B7E10). Southern, Northern and Western blotting analyses provide evidence for a lack of variability at the protein and DNA levels. A subclone of the insert in the expression vector pEX2, synthesises a fusion peptide which contains the epitope recognized by B7E10. Sequences homologous to the insert were detected in the genome of three other rodent and two primate malarias.

Animals↗

PubMed1988

Monoclonal antibodies to a synthetic peptide corresponding to a repeated sequence in the Plasmodium falciparum antigen Pf155.

Mouse monoclonal antibodies were prepared against a synthetic peptide (EENVEHDA) corresponding to a tandemly repeated sequence in the C-terminus of the Plasmodium falciparum antigen Pf155. One antibody (IgG1) producing hybridoma was studied in detail. The specificity of the antibody was determined by enzyme-linked immunosorbent assays using bovine serum albumin-conjugated or free peptides as solid phase antigens and various synthetic peptides for inhibition. The antibody reacted with Pf155 as detected by immunofluorescence and immunoblotting. It was also an efficient inhibitor of merozoite invasion in P. falciparum in vitro cultures indicating that it defines a biologically important epitope present on the native Pf155 molecule.

Animals↗

PubMed1988

Malaria vaccines: immunogen selection and epitope mapping.

In recent years major efforts have been made to characterize parasite antigens thought to be suitable candidates for malaria vaccines. Many of the relevant plasmodial antigens have been found to contain extensive areas of short amino acid sequences organized in tandem repeats. These are usually strongly antigenic, forming linear epitopes seen by antibodies of the infected host. Several such epitopes have been identified and subunit vaccines are being designed in which synthetic peptides or gene constructs serve as immunogens. However, as an efficient malaria vaccine should give rise to anamnestic T-dependent antibody responses following reinfection after vaccination as well as to antibody independent cell-mediated immunity, efforts are now also being made to identify T-cell epitopes on the vaccine candidate antigens. In this paper the current Plasmodium falciparum sporozoite vaccines and the merozoite antigen Pf155/RESA, a possible candidate for a P. falciparum blood stage vaccine, serve as examples to illustrate recent advances made in this area as well as some of the problems remaining to be resolved.

Animals↗

PubMed1988

T-cell epitopes in Pf155/RESA, a major candidate for a Plasmodium falciparum malaria vaccine.

Immunogens included in a subunit vaccine should contain both B- and T-cell-activating sites to ensure anamnestic responses following reinfection after vaccination as well as antibody-independent cellular immunity. The Plasmodium falciparum antigen Pf155/RESA, a major candidate for a vaccine against the asexual blood stages of this malaria parasite, was investigated for T-cell epitopes in its C-terminal amino acid repeat region, a region known to be conserved in different P. falciparum strains. It was found to contain several related sequences that activated T cells from humans primed to P. falciparum by natural exposure, to proliferation, and/or interferon-gamma release in vitro. T cells from approximately half of the donor group investigated responded to the intact protein, and 65% of these responders also responded to short synthetic peptides, probably representing a small number of partly overlapping T-cell epitopes. Thus, sequences from the C terminus of Pf155 may be suitable constituents of a P. falciparum subunit vaccine and also provide a basis for epitope-specific epidemiological studies relating cellular immune responses in vitro to clinical immunity and P. falciparum endemicity.

Amino Acid Sequence↗

PubMed1988

Induction of CD43 expression during activation and terminal differentiation of human B cells.

Only a small population (25-30%) of human peripheral blood B lymphocytes expresses large sialoglycoprotein (LSGP) (CD43). However, in the presence of autologous T cells and pokeweed mitogen (PWM) a majority (50-90%) of the immunoglobulin-producing cells (cIg+ cells) that develop from these B cells express CD43 is detected with anti-CD43 monoclonal antibodies (MoAb) B1B6, and the proportion of CD43+cIg+ cells increases with time of culture. Furthermore, a relatively larger proportion (60-80%) of the IgG-producing cIg+ cells are CD43+ compared with IgM-containing cIg+ cells (30-50%). In human tonsils, significantly more CD43+ cells (35%) are found in the in vivo-activated fraction of B cells than in the fraction of resting B cells (5%). A majority of the cIg+ cells that develop from the resting or the in vivo-activated tonsillar B cells in a PWM-induced B-cell differentiation system are CD43+ (80-100%). Furthermore, tonsillar B cells depleted of CD43+ cells give rise to cIg+ cells, of which the majority are CD43+, and the proportion of such cells increases with time of culture (60-90%). Taken together, these results indicate that LSGP belongs to a group of B-cell membrane molecules that are induced and upregulated upon activation and differentiation.

Antigens, CD↗

PubMed1988

Characterization of human CD4+ T-cell clones that secrete helper factor(s) for B-cell proliferation and maturation.

Human peripheral blood lymphocytes (PBL) were activated with K46M, a m mitogenic monoclonal antibody against La-reactive T lymphocyte surface structures. The cultures were expanded in the presence of interleukin 2 (IL-2). After 1 month of culture, the activated T cells were cloned by limiting dilution at 0.5 cells/well. Five clones with the CD3+CD4+ phenotype and one clone with the CD3+CD8+ phenotype were obtained. The CD3+CD8+ clone (K99) displayed a strong major histocompatibility complex (MHC)-unrestricted cytolytic activity against MOLT-4 and a weaker reactivity against the bladder tumour cell lines T24 and RT4. The natural killer (NK)-susceptible K562 cells were not lysed. Two of the CD3+CD4+ clones (K91 and K914) showed a helper activity in pokeweed mitogen (PWM)-induced IgG production by B cells. These cells differed in the expression of CD45R and CDw29 antigens, as defined by the monoclonal antibodies 2H4 or D10D11 and 4B4. When stimulated with PWM for 48 or 72 h, clone K91 and an additional CD4-positive clone (K913) secreted a factor into the supernatants which helped B cells to produce IgG. The K913 supernatant also induced some IgM production. The supernatant obtained after similar stimulation of K914 cells was inactive. None of these supernatants induced B cells to proliferate when tested together with phorbol myristate acetate (PMA). However, when K91 and K914 cells were activated with phytohaemagglutinin (PHA) for 48 or 72 h, the supernatant from K91 was strongly helpful in B-cell proliferation, whereas the supernatant from K914 cultures was only moderately active. In conclusion, we have established human T helper clones that release different factors supporting either B-cell proliferation or maturation when stimulated with PWM or PHA.

Antigens, CD↗

PubMed1988

Monoclonal antibodies against leucoagglutinin-reactive human T-lymphocyte surface components. I. Characterization of cellular binding sites.

The binding specificities of three biologically active anti-lymphocyte monoclonal antibodies (MoAb) (K46M, K3G, and 3-19-2) produced against human T-cell surface components reactive with the mitogenic lectin leucoagglutinin from Phaseolus vulgaris (La) were analysed. K46M is a strong T-cell mitogen, while K3G and 3-19-2 inhibited cell-mediated cytotoxicity. Resting peripheral blood lymphocytes (PBL) contained 4-16% K46M+ cells, 8-35% K3G+ cells, and less than 0.3-4% 3-19-2+ cells. After stimulation with T-cell mitogens the proportion of K46M+ and 3-19-2+ cells increased markedly (mean 59 and 30% positive cells, respectively), while the increase in K3G+ cells was less prominent (38%). K46M-reactive structures were expressed on mature T cells and probably also on B cells. K3G reacted with B and T cells while 3-19-2 showed a broader specificity reacting also with erythrocytes. All three MoAb reacted with lipid extracts of resting and activated PBL as well as with purified neutral glycolipids of lymphoid origin. In addition 3-19-2 reacted with lipid extracts of erythrocytes. K46M immuno-precipitated four surface peptides from lectin-stimulated PBL. Their apparent molecular weights were 53,000, 42,000, and 16,000 (doublet). The 53,000 and 42,000 MW peptides were identified as the alpha and beta chains of the T-cell antigen receptor. The identity of the 16,000 MW peptides is presently unknown. K3G and 3-19-2 did not specifically precipitate any lymphocyte surface peptide.

Animals↗

PubMed1988

Monoclonal antibodies against leucoagglutinin-reactive human T-lymphocyte surface components. II. Studies on the mechanism of K46M-induced activation and determination of the frequency of responding cells.

A monoclonal antibody, K46M (IgM kappa), obtained after immunization with leucoagglutinin (La)-reactive T-cell surface components, stimulated human lymphocytes to proliferate. It induced maximal proliferation at greater than 20 micrograms IgM/ml after 3-4 days of culture. Cells stimulated by K46M produced interleukin 2 (IL-2) and gamma interferon (IFN-gamma) and expressed receptors for IL-2 and transferrin. The majority of the activated cells were phenotypically T cells as defined by monoclonal antibodies against CD3 and CD2, and an increase in the K46M-positive cells was also observed during the activation period. K46M-activated cells display major histocompatibility complex (MHC)-unrestricted cytotoxicity against several cultured target cells. The frequencies of the cytotoxic and of the proliferative precursor cells were determined using a limiting dilution assay. K46M seems to activate a larger fraction of cytotoxic precursor cells against Molt 4 than against K562, but the statistical significance of these observations requires further exploration. Both K46M or La activated 40% of PBL to proliferate, whereas 70% of PBL were induced by OKT3. However, the frequency of K46M-activated cells was 40% only when the lymphocytes were plated at low cell densities, i.e. less than 0.5 cells per well. At higher densities an inhibition of proliferation was seen that resulted in a biphasic response curve, indicating that the activation of PBL by K46M was not a single hit event. This was not found with either La or OKT3. Whether K46M, in contrast to OKT3 and La, activates a subpopulation with suppressor activity remains to be established.

Animals↗