人参皂甙Rb1的免疫佐剂作用

分别将人参皂甙Rbl(人参主要成分之一)以及氢氧化铝胶作为佐荆和金黄色葡萄球菌菌体抗原混合免疫豚鼠，观察免疫前后血清抗体滴度变化；并用Rbl和奶牛乳房炎金黄色葡萄球菌疫苗混合免疫奶牛，观察免疫前后血清抗体滴度变化，以及血液淋巴细胞在刀豆蛋白A(ConA)、美洲商陆(PWM)和金黄色葡萄球菌抗原刺激下的体外细胞增殖反应。结果表明，Rbl和抗原混合物免疫动物后，未见任何不良反应；Rbl组豚鼠血清抗体滴度比对照组和氢氧化铝胶佐剂组滴度增加快，幅度显著增高；Rbl组奶牛血清抗体滴度比对照组奶牛显著增加，ConA、PWM和金黄色葡萄球菌抗原诱导的血液淋巴细胞体外细胞增殖反应比对照组显著提高。因此，Rbl是一种有效的免疫佐剂。     

Kinetics and type of immune response following intranasal and subcutaneous immunisation of calves

Protection of animals against respiratory infections has long been known to depend on respiratory mucosal immunity. However, few studies have been reported on the immune response following intranasal (i.n.) immunisation with non-living, soluble antigens. This study determined the kinetics of the humoral and cellular immune responses in calves after i.n. immunisation with Limulus haemocyanin (LH) with cholera toxin adjuvant, or subcutaneous (s.c.) immunisation with LH in incomplete Freund's adjuvant. A proliferative response of peripheral blood mononuclear cells cultured in vitro with LH was observed in animals immunised 7-10 days after i.n. and s.c. immunisations with no significant differences between the two immunised groups. LH -specific antibody was present in the serum of animals immunised s.c. (IgM, IgG1 and IgG2) and i.n. (IgA). Although significant IgA responses were observed, i.n. immunisations in cattle with soluble protein antigens and cholera toxin as an adjuvant did not induce a strong systemic immune response.     

Modulation of IgG subclass expression during antibody responses in sheep

Experiments were undertaken to investigate IgG subclass expression during epitope-specific antibody responses in sheep. Animals were immunised with conjugates of bovine serum albumin-DNP (BSA-DNP), or killed Staphylococcus aureus-DNP (Sa-DNP) alone or with muramyl dipeptide (MDP), dextran sulphate (DXS), or staphylococcal exotoxins (toxin). Sheep received two injections of the same preparation intracutaneously at six weeks interval. Total and IgG subclass-specific anti-DNP, and anti-carrier (BSA or S aureus) antibody levels were measured by ELISA in blood taken at weekly intervals before and after immunisation. Anti-DNP antibody levels in animals given Sa-DNP alone were considerably greater than in those immunised with BSA-DNP alone. Toxin suppressed antibody responses to DNP and both carriers; MDP suppressed anti-hapten antibody responses below the levels obtained with antigen alone. Neither toxin nor MDP significantly altered the IgG subclass profile of antibody to DNP bound to either carrier. DXS did not significantly change total levels of anti-DNP antibody measured in sheep given BSA-DNP or S aureus-DNP. However, DXS promoted IgG1 and suppressed IgG2 anti-DNP antibody responses during the secondary response to Sa-DNP but not to BSA-DNP.     

Development and duration of protection against salmonellosis in mice and sheep immunised with live aromatic-dependent Salmonella typhimurium.

The aim of this investigation was to determine the development and duration of protection in mice or sheep immunised with aromatic-dependent (aro-) Salmonella typhimurium strain CS332, by either parenteral or oral routes. Immunisation of mice by the intraperitoneal or sheep by the intramuscular routes was found to impart protection against oral challenge with the virulent parent S typhimurium strain CS94 as early as seven days after immunisation. In contrast, when immunisation was carried out by the oral route, protection was not evident until three weeks after immunisation. Regardless of the route of immunisation, mice were still partially protected at three months and were fully susceptible at six months after immunisation. In sheep, protection persisted for six months but not 12 months after immunisation. Only parenterally immunised mice and sheep developed high ELISA and, or, agglutinating antibody titres, and cutaneous delayed-type hypersensitivity (DTH) at three weeks after immunisation. Although both antibody and DTH were detectable three months after immunisation of mice with aro- S typhimurium strain CS332, none was detected at six months. Antibody measured by agglutination and ELISA was detectable six months after immunisation in sheep, although no DTH was evident. At 12 months after immunisation low levels of anti-LPS antibody (measurable by ELISA only) were detected in sheep immunised by the intramuscular route.     

Assessment of T-dependent and T-independent immune responses in cattle using a B cell ELISPOT assay

Understanding the mechanisms that maintain protective antibody levels after immunisation is important for vaccine design. In this study, we have determined the kinetics of plasma and memory B cells detectable in the blood of cattle immunised with model T-dependent or T-independent antigens. Immunisation with the T-D antigen resulted in an expansion of TNP-specific plasma cells post-TNP primary and booster immunisations, which was associated with increased titres of TNP-specific IgG antibodies. Although no TNP-specific memory B cells were detected in the T-D group following the primary immunisation, we detected an increase in the number of TNP-specific memory B cells post-TNP boost. In contrast, no TNP-specific plasma or memory B cells were detected after primary or secondary immunisation with the T-I antigen. We then investigated if immunisation with a third party antigen (tetanus toxin fragment C, TTC) would result in a bystander stimulation and increase the number of TNP-specific plasma and memory B cells in the T-D and/or T-I group. TTC immunisation in the T-D group resulted in a small increase in the number of TNP-specific plasma cells post-TTC primary immunisation and boost, and in an increase in the number of TNP-specific memory B cells post-TTC boost. This bystander effect was not observed in the animals previously immunised with the T-I antigen. In conclusion, the present study characterised for the first time the B cell response in cattle to immunisation with T-D and T-I antigens and showed that bystander stimulation of an established T-D B cell memory response may occur in cattle.     

Effect of corticosterone infusion on plasma corticosterone concentration, antibody production, circulating leukocytes and growth in chicken lines selected for humoral immune responsiveness

The effect of corticosterone on antibody production was studied in chicken lines selected for humoral immune response. 2. Twelve cockerels (33 days old) from lines selected for high or low antibody responses after immunisation with sheep red blood cells (SRBC) were implanted with mini-infusion pumps delivering corticosterone or vehicle continuously for 14 d. 3. Three days after implantation, the chickens were immunised intramuscularly with 0.25 ml packed SRBC. Blood samples were taken before implantation, before immunisation and 3, 5, 7 and 11 d after immunization. 4. Corticosterone infusion induced higher plasma corticosterone concentrations and heterophil/lymphocyte ratios than infusion of vehicle only. Growth was considerably depressed and relative weights of the thymus, bursa of Fabricius and spleen were less in the corticosterone-infused chickens. 5. An effect of corticosterone on antibody production could not be demonstrated, and differences between selection lines were unaffected.     

Humoral immunity in the ewe. 1. The influence of adjuvancy and immunogenic substitution on immune reactivity following immunisation with protein antigens

The nature of antigen and presence of adjuvant are major factors which influence the level of immune reactivity following immunisation. This study examines the quantity and isotype of antibody produced in ewes immunised with different proteins in combination with different adjuvants. Results using indirect ELISA assays show that animals immunised with keyhole limpet haemocyanin (KLH) in adjuvant produced lower levels (P less than 0.05) of immunoglobulin M (IgM) and had increased levels and persistence of immunoglobulin G1 (IgG1) compared with ewes immunised with antigen in saline (P less than 0.005). The anti-KLH immunoglobulin G2 (IgG2) titre was significantly higher in animals given oil-emulsion adjuvant than all other groups (P less than 0.005). Ewes were also immunised with bovine serum albumin (BSA) or BSA haptenated with trinitrophenyl (TNP) for the primary injection and carrier BSA alone for the secondary inoculation. Chemical haptenation of BSA antigen reduced primary IgM (P less than 0.005), IgG1 (P less than 0.005) and IgG2 (P less than 0.005) levels compared with animals immunised with pure BSA. There was an increased secondary anti-BSA IgM response in all animals first immunised with TNP-BSA (P less than 0.05). The class of anti-hapten antibody produced to TNP determinants was influenced by the degree of TNP haptenation of the carrier BSA. Mid-range molar ratios of TNP produced the strongest IgM, IgG1 and IgG2 anti-TNP responses compared with all other groups (P less than 0.01).     

Modulation of the immunogenicity of the Trypanosoma congolense cysteine protease, congopain, through complexation with ��2-macroglobulin

The protozoan parasite Trypanosoma congolense is the main causative agent of livestock trypanosomosis. Congopain, the major lysosomal cysteine proteinase of T. congolense, contributes to disease pathogenesis, and antibody-mediated inhibition of this enzyme may contribute to mechanisms of trypanotolerance. The potential of different adjuvants to facilitate the production of antibodies that would inhibit congopain activity was evaluated in the present study. Rabbits were immunised with the recombinant catalytic domain of congopain (C2), either without adjuvant, with Freund’s adjuvant or complexed with bovine or rabbit α₂-macroglobulin (α₂M). The antibodies were assessed for inhibition of congopain activity. Rabbits immunised with C2 alone produced barely detectable anti-C2 antibody levels and these antibodies had no effect on recombinant C2 or native congopain activity. Rabbits immunised with C2 and Freund’s adjuvant produced the highest levels of anti-C2 antibodies. These antibodies either inhibited C2 and native congopain activity to a small degree, or enhanced their activity, depending on time of production after initial immunisation. Rabbits receiving C2-α₂M complexes produced moderate levels of anti-C2 antibodies and these antibodies consistently showed the best inhibition of C2 and native congopain activity of all the antibodies, with maximum inhibition of 65%. Results of this study suggest that antibodies inhibiting congopain activity could be raised in livestock with a congopain catalytic domain-α₂M complex. This approach improves the effectiveness of the antigen as an anti-disease vaccine candidate for African trypanosomosis.     

Origin of antibody in porcine bile after intraperitoneal immunisation

Pigs were immunised intraperitoneally with ovalbumin (OVA) in Freund's complete adjuvant and killed between 52 and 71 days later. Sera, bile and spleen and liver tissue were collected at slaughter. IgG and IgA OVA antibody in bile and serum were detected by ELISA, and IgG and IgA anti-OVA containing cells (AOCC) in tissue were observed using double fluorochrome labelling techniques. The results indicated few IgA AOCC in spleen or liver, but an elevated IgA OVA response in bile compared to serum relative to IgG. The results indicate selective transport of IgA OVA antibody from serum to bile relative to IgG.     

Immunisation against lactic acidosis in cattle.

The present study was designed to investigate the efficacy of control of lactic acidosis by immunisation against lactic acid-producing bacteria, Streptococcus bovis and Lactobacillus. Ten steers were allocated to two treatment groups. One group was immunised with a vaccine containing S. bovis (strain Sb-5) and Lactobacillus (LB-27) cells, and the other was a non-immunised control group. The vaccine, using Freund's complete adjuvant for primary immunisation and Freund's incomplete adjuvant for boosters, was administered intramuscularly. After primary immunisation, boosters were given at 2 to 4 week intervals. Both anti- S. bovis and anti- Lactobacillus IgG levels in saliva increased significantly (P < 0.01) after the 1st booster which were lower (P < 0.05) than the IgG levels after the 2nd and 3rd boosters, but were not significantly different (P > 0.05) from the IgG levels prior to a grain challenge (after the 4th booster). There were positive correlations between the anti- S.bovis and anti- Lactobacillus IgG in serum and saliva. Compared with the control group, steers in the immunised group had higher (P < 0.05) feed intakes, lower (P < 0.05) rumen concentrations of lactate and lower numbers of S. bovis and Lactobacillus. Three of the control animals were withdrawn from the grain challenge due to their rumen pH persisting below 5.2, while only one animal in the immunised group was withdrawn. These results suggest that the risk of lactic acidosis can be reduced by immunisation against S. bovis and Lactobacillus.     

Blood lymphocyte subsets in pigs vaccinated and challenged with Actinobacillus pleuropneumoniae

Actinobacillus pleuropneumoniae bacterins do not induce protection in pigs while infection with low doses of the CM5 strain of A. pleuropneumoniae given by aerosol induces complete protection. To evaluate possible correlates of protection in blood lymphocyte subset phenotypes, pigs were treated with a commercial bacterin given intramuscularly, low dose (10(5)cfu/ml) aerosol infection with CM5 or control treatments of the bacterin adjuvant or phosphate buffered saline. All pigs were challenged with a high dose (10(7)cfu/ml) of A. pleuropneumoniae. Lymphocytes and sera were collected prior to and following primary and secondary immunizations and challenge, for evaluation of B- and T-cell markers and antibody to four A. pleuropneumoniae antigens. IgM(micro)+ B-cells were increased following primary exposure to antigen in the bacterin-vaccinated group only. An increase in CD4+ cells in the LD aerosol-infected group was apparent following secondary exposure to antigen. These early changes suggest little difference in lymphocyte populations between treatment groups, however, greater differences were observed following high-dose challenge; CD4+ lymphocytes were increased significantly in both bacterin and LD-challenged groups (p<0.05) while CD8+ cells decreased in the LD-group at this time period. Consequently, there were significant differences (p<0.05) in the CD4:CD8 ratio after high-dose challenge compared to earlier time points and control groups. Variation in cellular expression of SLA-DR and DQ was observed but trends correlating to treatment group were not evident. Complete protection or lack of protection associated with LD challenge or immunisation resulted in significant differences in B-cell frequencies and CD4:CD8 ratio phenotypes in pigs, but only changes in CD4:CD8 ratios appeared relevant to protection.     

Foetal cross-protection experiments between type 1 and type 2 bovine viral diarrhoea virus in pregnant ewes

A flock of 82 non-pregnant ewes was split into three immunisation groups and given an intranasal dose of either cell culture medium, or a type 1 or a type 2 bovine viral diarrhoea virus (BVDV-1 or BVDV-2). Two months later the flock was reconstituted and after a further three weeks, the ewes were bred to pestivirus negative rams after synchronisation of oestrus using progesterone sponges. Fifty-five ewes were segregated into three challenge groups, each of which comprised ewes from different immunisation groups. At 7 weeks gestation, one challenge group was given an intranasal dose of cell culture medium, whilst the other two were given intranasal doses of either BVDV-1 or BVDV-2, using the same inocula as for the immunisations. Three weeks later, the ewes were killed and their foetuses tested for the presence of BVDV-1 and BVDV-2. The results showed that immunisation of six ewes without subsequent challenge did not lead to infection of any of their 11 foetuses. Challenge with BVDV-1 or BVDV-2 in the absence of immunisation lead to 15 out of 15 or 11 out of 14 foetuses becoming infected, respectively. Immunisation with the homologous virus to that used for challenge resulted in complete protection of 32 foetuses from 15 ewes. Heterologous protection was one way. All 12 foetuses from ewes immunised with BVDV-1 were protected from challenge with BVDV-2, whereas 18 foetuses from ewes immunised with BVDV-2 were all infected after challenge with BVDV-1. This provides evidence that a recent exposure to infection with one pestivirus does not necessarily induce foetal protection against another. The one-way result suggests that factors other than antigenic differences are involved in cross-protection.     

Immunisation of chickens to reduce intestinal colonisation with Campylobacter jejuni

1. Systemic and intestinal antibody titres were measured in chickens following subcutaneous, intraperitoneal (IP), oral (po) and combined IP/po administration of antigen, in soluble, emulsified or microparticulate form. Antigens tested included keyhole limpet haemocyanin (KLH), killed Campylobacter jejuni whole cells and purified campylobacter flagellin protein.

2. The effect of immunisation with purified flagellin protein or with killed C. jejuni whole cells in reducing intestinal colonisation was assessed. The ability of newly‐hatched chicks to respond to immunisation was limited, possibly because of the immaturity of the immune system rather than maternal suppression of an immune response. Only 5 of 13 birds that were first immunised when 1‐d‐old with KLH showed a systemic response, even after 4 immunisations, whereas 10 of 11 birds that were first immunised at 24 d‐old responded systemically.

3. In an immunisation and challenge experiment, birds that were immunised twice intraperitoneally, at 16 and 29 d‐old, with killed C. jejuni whole cells, had fewer C. jejuni, in the caecal contents than unimmunised control birds. This reduction in intestinal colonisation, to less than 2% of bacterial numbers in control birds, was associated with an increase in specific IgG in intestinal secretions. There was no significant increase in specific IgA or IgM in intestinal secretions following immunisation and challenge.

4. These results indicate that immunisation can reduce the level of intestinal infection with C. jejuni. The protection may be enhanced by developing improved methods of immunisation that stimulate production of increased titres of specific antibody in intestinal secretions, particularly specific IgA antibody.     

The antibody response to bovine lymphocyte alloantigens

Antibodies were raised against lymphocyte cell-surface antigens by multiple immunisations with purified lymphocytes or by the exchange of skin allografts. Eighteen of 21 cattle immunised with lymphocytes raised a detectable cytotoxic antibody response. The serum antibody from 10 responders recognized only common lymphocyte antigens, those antigens which are present on all peripheral blood lymphocytes. One animal responded only to B lymphocyte antigens while 7 others responded to both classes of antigens. The amount of antibody produced varied greatly between individuals; antibody titres ranged from 1 to 1028. Antibody raised early in the response was sensitive to treatment with 2-mercaptoethanol (2-ME) suggesting that IgM was the predominant class of immunoglobulin. Subsequently antibody became resistant to this treatment suggesting the appearance of IgG. The antibody responses following the exchange of skin grafts were very similar in all 12 cattle studied. High titred antibody to common lymphocyte antigens was detected in the serum 14 days after grafting. The early antibody activity was sensitive to 2-ME treatment but became totally resistant within 14 days. Total peak antibody titres ranged from 128-2048. Antibody to B lymphocyte antigens was identified in 8 of the 12 cattle. The responses to B lymphocyte antigens were similar to those against the more widely distributed common lymphocyte antigens with respect to time of antibody appearance, time of peak titre and sensitivity to 2-ME. Peak titres ranged from 2 to 32. The change in antibody specificity with time was also studied. Sera from 11 of the 18 cattle which had responded against lymphocytes showed an increase or broadening in reaction frequency as immunisations increased, suggesting the production of antibody to secondary specificities. In the cattle which had been skin grafted, the broadest reaction patterns were seen 14 to 21 days after grafting. The broadest reaction patterns were seen when the antibody responses were at their highest titre levels and narrowed as titres decreased.     

Production of antibodies to recombinant antigens from Lucilia cuprina following cutaneous immunisation of sheep

Immunological control of cutaneous myiasis of sheep caused by Lucilia cuprina larvae has been an elusive goal. Antibody to antigens derived from the peritrophic membrane can stunt or kill larvae in a dose dependent fashion. Thus efficacy of vaccines employing these antigens may be limited by the amount of antibody in skin available for ingestion by larvae. The potential for elevating antibody concentrations in skin by intradermal immunisation with the recombinant peritrophic membrane antigens peritrophin-44, peritrophin-48 and peritrophin-95 was therefore examined. Using within-animal comparisons, specific antibody was significantly higher in skin transudates from locally immunised sites than from adjacent adjuvant control sites. It was concluded that cutaneous immunisation may assist immunological control of blowfly larvae.     

Humoral immunity in the ewe. 2. The effect of pregnancy on the primary and secondary antibody response to protein antigen

This study examines the effect of pregnancy on the quantity and isotype of antibody in ewes immunised with a novel primary antigen. The primary and secondary antibody levels to bovine serum albumin (BSA) in alum adjuvant, were compared between non-pregnant ewes and ewes immunised at different stages of pregnancy. Anti-BSA isotype specific responses were measured using an indirect ELISA. Results show the levels of immunoglobulin M (IgM) increased and persisted in response to BSA during pregnancy (P less than 0.05). Secondary immunoglobulin G1 (IgG1) titres were significantly impaired in late pregnancy and during lactation (P less than 0.05). The lower levels of immunoglobulin G2 (IgG2) were unaffected by pregnancy under these experimental conditions. Ewes were also immunised with BSA in alum adjuvant for their primary inoculum and BSA in different adjuvants for the secondary inoculation. Primary IgM persisted at higher levels during pregnancy compared with the response in non-pregnant ewes (P less than 0.05). Following the secondary injection, lower levels of anti-BSA specific IgM, IgG1 and IgG2 antibodies were produced in late pregnancy and during lactation compared with the levels in non-pregnant control ewes (P less than 0.05). Alterations in regulatory T-cell function or effector B-cell activity would most readily explain the qualitative changes in antibody titre observed following primary injection during pregnancy. The results also suggest an associated impairment of immunological memory following primary immunisation during pregnancy.     

Immune responsiveness and lymphoreticular morphology in cattle fed hypo- and hyperalimentative diets

Immune responses and lymphoreticular morphology were studied in 3 groups of yearling Hereford steers fed hypoalimentative, maintenance and hyperalimentative diets for 142 days. Significant decreases in plasma protein levels and circulating lymphocyte levels occurred in low energy intake steers. Percent circulating lymphocytes bearing surface immunoglobulins and serum levels of IgG and IgM did not vary significantly within or between groups. Antibody responses to Brucella abortus bacterin inoculated on day 63 were similar in all 3 groups. Antibody responses to chicken erythrocytes inoculated on day 88 were significantly lower in hypoalimentated steers. Differences between groups in lymphocyte blastogenic responses to phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM) were not significant. Hyperalimentated steers had significantly depressed PWM responses compared to a baseline value established for that group. In addition, hypoalimentated steers tended to have elevated responses to PHA, although differences were not significant. There were no significant differences between groups in dermal hypersensitivity responses to tuberculin following sensitization with viable Bacillus Calmette-Guerin (BCG). The thymuses of hypoalimentated steers were markedly atrophied but lymph nodes and splenic white pulp were normal. Thymus, lymph nodes and spleen were normal in hyperalimentated steers.     

Serum bactericidal responses to Streptococcus equi of horses following infection or vaccination

An indirect test based on horse blood was used to study bactericidal responses of the horse to Streptococcus equi following infection or vaccination. Bactericidal antibody appeared in convalescent sera between two and four weeks and high titres were usually attained by eight weeks. Infection without clinical evidence of abscessation was also effective in eliciting strong bactericidal responses. Serum bactericidal activity of horses either recovered from strangles or immunised with commercial bacterin had declined eight months after vaccination. However, horses that developed strangles eight to 10 months after vaccination exhibited rapid and substantial increases in serum bactericidal activity. Groups of yearlings immunised with commercial S equi vaccines consisting either of M protein or bacterin developed clinical strangles within six months of vaccination although the majority of the animals had exhibited strong serum bactericidal activity a few weeks before occurrence of the disease. Similarly, a group of seven yearling ponies hyperimmunised with experimental vaccine, rich in M protein, were found to be highly susceptible to an intranasal challenge of 5 X 10(8) colony forming units of S equi, although their sera exhibited strong bactericidal activity at the time of challenge. These observations suggest that the role of serum bactericidal antibody in protection of the horse against strangles has been overrated.     

Effects on adhesion molecule expression and lymphocytes in the bovine mammary gland following intra-mammary immunisation

Changes to adhesion molecule expression and lymphocyte populations were evaluated in alveolar mammary tissue collected from cows following an immunisation protocol that involved intra-mammary inoculation to induce an IgA response in mammary secretions. The right quarters of the udder were immunised; the left side acted as a control. Antibody titres in secretions showed that at least two animals responded with antigen-specific IgA. Numbers of T-lymphocytes were 4-fold higher in immunised glands compared with controls (P < 0.05). IgA-, IgM- and IgG-positive cell numbers were significantly higher (P < 0.01) in immunised glands compared with controls in three of the four cows. No mucosal addressin molecule-1 (MAdCAM-1), vascular cell-adhesion molecule-1 (VCAM-1) or peripheral node addressin (PNAd) protein expression was detected on smaller venules that stained positively for von Willebrand factor in alveolar mammary tissues, from either immunised or control glands. Both VCAM-1 and PNAd were detected on smaller venules in supramammary lymph nodes, however, there was no significant difference between immunised and control glands. Quantification of MAdCAM-1 mRNA showed very low expression in both immunised and control alveolar tissue compared with Peyer's patch positive-control tissue. These findings suggest that the bovine mammary gland is capable of a mucosal antibody response; however, MAdCAM-1 is not involved with lymphocyte homing to the mammary gland in this species.     

Characterization of monoclonal antibodies directed against swine leukocytes

Hybridomas were produced from fusions of the SP2/0 mouse myeloma with splenic cells from: 1) an outbred Sprague Dawley rat immunized with swine peripheral blood mononuclear (PBM) cells; 2) a (CBA/NDub X BALB/c Dub) F1 mouse immunized with concanavalin A (Con A) activated swine PBM cells and 3) a (BALB/c Dub X C3H/He Dub) F1 mouse immunized with swine thymocytes. The resulting supernatants were screened by a microcytotoxicity assay for activity against swine PBM cells. Four hybridomas (MSA1, MSA2, MSA3 and MSA4) were selected, cloned and characterized by their cell reactivity and effect on mitogenic assays. MSA1 and MSA2 belong to the rat IgG2b subclass. MSA3 and MSA4 are of the mouse IgG2a subclass. These monoclonal antibodies reacted in the following manner: MSA1 with monocytes, granulocytes, red blood cells and bone marrow cells; MSA2 with subset of T cells; MSA3 with B cells and subsets of T cells and monocytes (class II molecule) and MSA4, a pan-T cell reagent (E-rosette receptor). The involvement of the various cell types reactive to the different monoclonal antibodies in the mitogenic response of swine PBM cells to Con A, phytohemagglutinin (PHA) or pokeweed mitogen (PWM) was investigated by cellular depletion with monoclonal antibody plus complement. Cellular depletion of PBM cells with the following monoclonal antibodies plus complement treatment resulted in: MSA1, almost total reduction in the mitogenic response to low doses of Con A or PWM; MSA2, partial reduction in the proliferative responses to any concentration of Con A, PHA or PWM; MSA3, partial reduction in proliferative responses to low concentrations of Con A or PWM and 4) MSA4, total elimination of any proliferative response to Con A, PHA or PWM.