Analysis of IgG subclass antibodies and expression of T-Cell receptor signaling molecules in anti-CD4 monoclonal antibody treated mice with autoimmune cardiomyopathy

T-cell immune abnormality in patients of dilated cardiomyopathy has been intensively studied over the past 10 years. In this study, we aim to focus on the molecular mechanism of T-cells in autoimmune cardiomyopathy mouse model by detecting the expression of three T-cell signaling molecules. Balb/C mice (n = 12) were immunized with the peptides derived from human ADP/ATP carrier on the 1st, 14th, 28th, 49th and 79th days, and half of them were also injected with anti-L3T4 McAb on the - 1st, 0 and 1st days. The sham-immunized mice were taken as the controls (n = 6). The main result shows that the antibody response of IgG subclasses such as IgG1, IgG2b and IgG3 were definitely blocked except IgG2a in CD4+ cell-depleted Balb/C mice. In addition, the average mRNA expression of p56lck, p59fyn and zap-70 were all found to be dramatically higher in the mice immunized with only ADP/ATP carrier peptides than in the control-group. At meantime, reduced levels of the protein kinases p56lck, p59fyn and zap-70 were clearly observed in anti-CD4 McAb immunized group compared with DCM group. We propose that the proliferation of T-cells was significantly inhibited in anti-CD4 treated mice and CD4+ T-cells may play a critical role in ADP/ATP carrier caused mouse DCM.     

Infectious Tolerance to ADP/ATP Carrier Peptides Induced by Anti-L<Subscript>3</Subscript>T<Subscript>4</Subscript> Monoclonal Antibody in Dilated Cardiomyopathy Mice

CD4 T cells are suspected to play an important role in the pathogenesis of dilated cardiomyopathy (DCM). This study sought to evaluate whether anti-L3T4 monoclonal antibody (McAb) could induce the infectious tolerance to the adenosine diphosphate (ADP)/adenosine triphosphate (ATP) carrier peptides to protect mice from DCM. BALB/c mice (n = 16) were immunized with the peptides derived from human ADP/ATP carrier on the 1st, 14th, 28th, 49{th}, and 79th days, and some of them (n = 6) were also injected with anti-L₃T₄ McAb on the −1{st}, 0, and 1st days. On the 180th day, the splenocytes (SC) from the McAb-treated group were transferred into the syngeneic recipients (n = 6) who were also immunized with the peptides in the same manner. The sham-immunized mice were taken as the controls (n = 10). Results showed that the serum antibody against the ADP/ATP carrier examined with ELISA was positive in all mice only immunized with the peptides (DCM group), while negative in the McAb-treated, the SC-transferred, and the Control groups. The mRNA expression of IFN-γ, IL-2, and IL-4, especially IL-4 in T cells investigated using real-time quantitative PCR and the percentages of T helper 1 (Th1) and Th2 subsets, especially Th2 subset detected with Flow Cytometry were all increased in DCM group, accompanied by the cardiac histopathological changes like those in DCM. Such findings were not seen in the other three groups. It concluded that anti-L₃T₄ McAb could inhibit the occurrence of DCM induced by the ADP/ATP carrier peptides in mice, and this immune tolerance could be transferred to the syngeneic recipients.     

抗CD4单抗干预实验鼠自身免疫性心肌病的T细胞信号转导机制(英文)

目的：探讨抗L3T4单克隆抗体对BALB/c小鼠实验性自身免疫性心肌病的治疗作用及T细胞信号转导机制。方法：以ADP/ATP载体肽免疫小鼠建立自身免疫性心肌病模型。3个月后，向小鼠体内注入抗L3T4单抗，以清除CD4+T细胞，并以无关抗体为对照。通过实时荧光定量PCR检测小鼠T细胞中信号分子（p56lck、p59fyn和Zap-70）和细胞因子（IFN-γ、IL-2和IL-4）的表达；通过免疫组化技术分析各组小鼠T细胞内CD45的表达。结果：给予抗L3T4单抗治疗的DCM小鼠T细胞内信号分子（p56lck、p59fyn和Zap-70）和细胞因子（IFN-γ、IL-2和IL-4）表达均减少，CD45的表达也明显减少。相反，以无关抗体免疫小鼠并不能对其产生保护作用，T细胞信号分子、CD45和细胞因子的表达也不被抑制。结论：抗L3T4单抗可以抑制ADP/ATP载体肽诱导的小鼠DCM过程中的T细胞信号分子和细胞因子的表达，可能起到治疗作用。     

Treatment with an anti-CD4 monoclonal antibody strongly ameliorates established rat adjuvant arthritis

Some experimental arthritic diseases can be prevented by treatment with anti-CD4 MoAbs. Trials with ongoing disease have not been successful so far. The aim of this study was to ascertain whether W3/25 could reverse adjuvant arthritis (AA), when beginning treatment on day 14, i.e. when the disease was established. Moreover, one group of animals treated with the anti-CD4 MoAb received OX8 MoAb at the same time, thus depleting CD8⁺ cells from circulation. During treatment with W3/25, a strong amelioration of inflammatory signals was observed, as assessed by means of paw volume increase and arthritic score. However, when treatment stopped, a rebound to arthritis signals occurred. The parallel depletion of CD8⁺ cells did not modify these effects, thus the combined treatment W3/25+OX8 gave the same amelioration as treatment with W3/25 alone. These findings indicate that CD4⁺ cells play an important role in perpetuating rat AA. Moreover, CD8⁺ cells do not seem to have a regulatory role in the CD4⁺ cells responsible for the inflammatory response.     

不同阶段应用抗L3T4单抗干预治疗对心肌病小鼠细胞因子的影响

目的： 研究不同阶段应用抗L3T4单抗对心肌病小鼠Th1/Th2亚群及血清和心肌组织中细胞因子的影响，探讨抗L3T4单抗治疗自身免疫性心肌病的机制。方法: 用含有人线粒体ADP/ATP载体肽的免疫液免疫近交系BALB/c小鼠建立类扩张型心肌病模型（心肌病组）；以不含肽免疫液免疫小鼠作为对照组；在用ADP/ATP载体肽免疫小鼠的前1 d连续3 d以400 μg抗L3T4单抗免疫小鼠获得早期治疗组；心肌病组小鼠于第4个月初始连续3 d给予抗L3T4单抗治疗获得中期治疗组，单抗使用方法同早期治疗组。运用3色荧光标记流式细胞术检测小鼠脾脏中Th1/Th2的百分含量；以ELISA法检测其血清中IFN-γ、IL-2、IL-4、IL-6和TNF-α水平；实时荧光定量PCR法检测其心肌细胞因子基因表达。 结果: 早期治疗组Th1及Th2亚群明显低于心肌病组；中期治疗组Th1相关细胞因子水平高于心肌病组，Th2水平介于心肌病组和早期治疗组之间。早期治疗组IFN-γ和IL-6与对照组相近，IL-2和TNF-α均高于对照组和心肌病组，IL-4介于前两组之间且与它们均有显著差异；中期治疗组IFN-γ和IL-2水平介于对照组和心肌病组之间，IL-6和IL-4明显低于心肌病组。 结论: 不同阶段应用抗L3T4单抗能够阻断或减轻系统性和局部细胞因子的生成，早期治疗较中期治疗对细胞因子的抑制作用更显著。     

Lymphocyte changes associated with prolongation of cardiac allograft survival in adult mice using anti-CD4 monoclonal antibody.

This study investigated the effect of anti-CD4 MoAb treatment on lymphocyte phenotype and function and correlated these changes with the prolongation of cardiac allograft survival in adult mice. Indefinite survival of heterotopic cardiac allografts was obtained in several fully allogeneic strain combinations when two doses of the anti-CD4 MoAb, YTS 191.1, were given at the time of transplantation. A dose response analysis in the C57BL/10 to C3H/He strain combination showed that very low doses of YTS 191.1 (25 micrograms/dose) were able to induce prolonged allograft survival when administered perioperatively. At the time of transplantation the immunosuppression induced by administration of the anti-CD4 MoAb is not antigen-specific, as heart grafts from different donor strains, mismatched for both major and minor histocompatibility antigens, showed prolonged survival in treated recipients. Immunocompetence was restored by 6 weeks after MoAb treatment, as recipients regained the ability to reject a cardiac allograft transplanted at this time point. However, while recovery of immunocompetence could be demonstrated in vivo, leucocytes isolated from the peripheral lymphoid organs of treated mice continued to be hyporesponsive in mixed leucocyte culture (MLC). Phenotypic analysis of the peripheral lymphoid tissues showed that C3H/He recipients treated with 25 micrograms/dose of YTS 191.1 had a marked, but not complete, elimination of the CD4+ subset at the time of transplantation, which was gradually restored to 50% of normal by 6 weeks after treatment. Thus, complete elimination of the CD4+ subset was not required to achieve indefinite allograft survival, and immunocompetence, as assessed in vivo, returned even when the CD4+ subset was present at half the normal level. Low doses of anti-CD4 MoAb (25 micrograms) had no effect on the expression of the CD4 molecule by thymocytes, and yet thymocytes were hyporesponsive to alloantigen in vitro. At higher doses of YTS 191.1, immature CD4+8+ thymocytes were selectively depleted. These results suggest that anti-CD4 MoAb therapy may modulate the intrathymic T cell selection process. These studies provide further insight into the mechanism of action of low dose, depleting anti-CD4 MoAb therapy in allograft rejection, and form a basis from which rational modifications to therapeutic protocols in transplantation models can be made.     

Rat brain xenografts reverse hypogonadism in mice immunosuppressed with anti-CD4 monoclonal antibody

Summary This study examines the effect of immunosuppression with monoclonal antibodies (MAb) against the murine CD4 (L3T4), a cell surface glycoprotein expressed primarily on helper T-lymphocytes, on the viability and function of rat neural xenografts placed in the third ventricle of hypogonadal (hpg) mice. The hpg mouse fails to synthesize hypothalamic gonadotrophin releasing hormone (GnRH) and consequently there is a drastic reduction in pituitary gonadotrophic hormone content and a failure of postnatal gonadal development (Cattanach et al. 1977). Three groups of male hpg mice received xenografts of day 1 post natal rat preoptic area (POA) tissue, a source of GnRH neurons, to their third ventricle. Those immunosuppressed with anti-CD4 MAb all showed surviving graft tissue thirty days post-transplant and half of this group had enlarged testes with all stages of spermatogenesis. In those hpg mice which were injected with saline alone, or with an anti-CD8 (Lyt-2) antibody there was no xenograft survival. These results suggest that the injection of monoclonal antibodies against the T-helper subset may provide an alternative means of immunosuppression aimed at the enhancement of survival of tissue grafts in the CNS.     

Chimaeric anti-CD4 monoclonal antibody cross-linked by monocyte Fcγ receptor mediates apoptosis of human CD4 lymphocytes

Previous studies have shown that murine anti-CD4 monoclonal antibody, cross-linked by rabbit anti-mouse immunoglobulin, could mediate apoptosis of murine CD4⁺ lymphocytes when they were stimulated by T cell receptor antibody. In this study, we have shown that the murine anti-CD4 monoclonal antibody, OKT4, can induce apoptosis in human CD4⁺ T cells stimulated by the recall antigen tuberculin purified protein derivative (PPD) only when cross-linked by rabbit anti-mouse immunoglobulin. The chimeric anti-CD4 monoclonal antibody, cM-T412 whose Fc fragment is human, was able to cause apoptosis without cross-linking by a second antibody. Similarly, abolition of PPD-induced proliferation of peripheral blood mononuclear cells by cM-T412 did not require cross-linking with rabbit anti-human immunoglobulin. Inhibition of proliferation by cM-T412 could be reduced by pre-treating monocytes with heat-aggregated human IgG. This suggested that monocyte Fcγ receptors might be cross-linking the human Fc of cM-T412. Propidium iodide staining together with immunofluorescence showed that the apoptotic cells were indeed CD4⁺ lymphocytes. It is proposed that during treatment with cM-T412 in autoimmune disease such as rheumatoid arthritis, cM-T412-coated CD4 T cells, when they are subsequently stimulated by the unknown arthritogenic antigen, may undergo apoptotic cell death through cross-linking of cM-T412 on Fey receptor-positive cells within the joint.     

In vivo treatment with a monoclonal chimeric anti-CD4 antibody results in prolonged depletion of circulating CD4+ cells in chimpanzees

Chimeric M-T412 (cM-T412), an anti-CD4 antibody, was tolerated in chimpanzees at a dosage of 5 mg/kg per day for up to 7 consecutive days, or 5 mg/kg per dose, twice weekly for 4 weeks. All cM-T412-treated chimpanzees showed a prolonged CD4 cell depression. Weak chimpanzee antibody responses to chimeric M-T412 were observed. One of the chimpanzees on the biweekly dosage regimen exhibited a hypersensitivity reaction immediately after receiving its seventh dose. Following supportive treatment, the animal recovered and remained asymptomatic during the non-treatment observation period. The hypersensitivity reaction was not an unexpected response considering the animal received repeated intermittent i.v. administration of a foreign protein. This animal also showed a chimpanzee antibody response to chimeric M-T412 after the seventh dose. Chimeric M-T412 also induced an anti-cM-T412 response in some of the other animals. The level of this response was lower than the anti-mouse responses observed in animals treated with murine anti-CD4. Moreover, the anti-cM-T412 response was mainly directed to idiotypic determinants. The decrease in CD4⁺ cells observed for all chimeric M-T412-treated chimpanzees is an expected effect of the anti-CD4 antibody. The duration of this CD4⁺ cell decrease is, however, much longer than observed for other CD4-specific MoAbs described. No selective loss of either memory or naive CD4⁺ cells was observed after either the single, 7-day or twice-weekly treatments. The CD4⁺ cell depression was reversible, although individual variation in time to recovery was observed. Therefore, cM-T412 could be a good candidate for clinical use in autoimmune conditions.     

Prevention of adjuvant arthritis by the W3/25 anti-CD4 monoclonal antibody is associated with a decrease of blood CD4(+)CD45RC(high) T cells

Imbalance between Th1 and Th2 functions is considered to play a key role in the induction and development of several autoimmune diseases, and the correction of that imbalance has led to effective therapies of some experimental pathologies. To examine whether CD4(+)CD45RC(high) (Th1-like) and CD4(+)CD45RC(low) (Th2-like) lymphocytes play a role in the pathogenesis of adjuvant arthritis (AA) and in its prevention by anti-CD4 antibody, CD45RC expression on CD4(+) T cells was determined in arthritic rats and in animals treated with an anti-CD4 MoAb (W3/25) during the latency period of AA. The phenotype of regional lymph node lymphocytes from arthritic rats in the active phase of the disease was determined by flow cytometry. Peripheral blood lymphocytes from rats treated with W3/25 MoAb were also analysed for 2 weeks after immunotherapy finished. IgG2a and IgG1 isotypes of sera antibodies against the AA-inducing mycobacteria, considered to be associated with Th1 and Th2 responses, respectively, were also determined by ELISA techniques. Fourteen days after arthritis induction, regional lymph nodes presented an increase in CD4+CD45RC(high) T cell proportion. Preventive immunotherapy with W3/25 MoAb inhibited the external signs of arthritis and produced a specific decrease in blood CD4(+)CD45RC(high) T cells and a diminution of antibodies against mycobacteria, more marked for IgG2a than for IgG1 isotype. These results indicate a possible role of CD4(+)CD45RC(high) T lymphocytes in the pathogenesis of AA, and suggest that the success of anti-CD4 treatment is due to a specific effect on CD4(+)CD45RC(high) T subset that could be associated with a decrease in Th1 activity.     

重组牛白细胞介素4的原核表达及其单克隆抗体研制

目的:原核表达重组牛白细胞介素4(rBoIL-4)并制备针对抗rBolL-4的单克隆抗体(mAb).方法:通过PCR从重组质粒pSP73-BOIL-4中扩增BoIFN-γ基因,分别克隆入表达载体pGEX-6p-1和pET-30a(+)中,构建重组菌并对其进行诱导表达、纯化和鉴定.以纯化的融合蛋白rHis-BoIL-4为...     

抗CD25单克隆抗体对CD4+CD25high调节性T细胞的影响

目的 评价抗CD25单克隆抗体对活体肾移植受者外周血CD4 CD25high调节性T细胞(Treg)的影响.方法 观察14例活体肾移植受者外周血CD4 CD25high T细胞百分比和FoxP3mRNA表达水平的变化,应用流式细胞仪FACS Calibur和CellQuest软件行双色流式细胞计数和结果分析,半定量逆转录-聚合酶链反应(RT-PCR)对肾移植受者外周血单个核细胞(MNC)的FoxP3基因水平进行检测.结果 在移植术后3d、13d,CD4 CD25high T细胞占CD4 比例显著升高;在移植后1个月内,CD4 CD25high T细胞比例升高约至原来的3倍.移植术后13d、17d、27d时外周血FoxP3 mRNA表达明显高于移植术后3d.在肾移植术后的不同时间抗CD25单克隆抗体使用组受者的CD25 T细胞百分比明显低于对照组(未使用抗CD25单克隆抗体).抗CD25单克隆抗体使用组与对照组两组间比较,术后3d、13d,抗CD25单克隆抗体组CD4 CD25high T细胞占CD4 的比例明显降低,术后3d对照组、抗CD25单克隆抗体使用组分别为1.24%±0.15%、2.25%±0.24%,P=0.00;术后13d为3.75%±0.38%、7.28%±0.51%,P=0.00.然而,术后17d、27d后,抗CD25单克隆抗体使用组的CD4 CD25high T恢复到对照组水平,两组间CD4 CD25high T细胞占CD4 的比例无统计学差异,即第17d为3.72%、0.26%、4.43%、0.44%,P=0.17,27d为9.00%±0.30%、8.31%±0.33%,P=0.16.在肾移植术后不同时间,受者外周血FoxP3mRNA表达水平在两组之间相似,无统计学差异,说明FoxP3mRNA表达水平并没有受到体内注射抗CD25单克隆抗体影响.结论 抗CD25单克隆抗体的使用对活体肾移植受者外周血CD4 CD25high调节性T细胞在一定时间具有暂时性的影响,而FoxP3 mRNA表达水平没有受到影响.     

Elisa test system with monoclonal antibodies to human IgG4 for the detection of allergen-specific antibodies in pollinosis

G. N. Gabrichevskii Moscow Research Institute of Epidemiology and Microbiology. Institute. of Experimental Cardiology, All-Union Cardiologic Scientific Center, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. I. Vorob'ev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 108, No. 11, pp. 574–577, November, 1989.     

Monoclonal antibodies to epitope of CD45R (B220) inhibit interleukin 4-mediated B cell proliferation and differentiation.

A mAb, ALP-2, produced by immunizing rats with proliferating lymph node cells from MRL/Mp-lpr/lpr mice, had an inhibitory effect on recombinant interleukin 4 (rIL-4)-mediated proliferation and differentiation of murine B cells. Kinetic analysis revealed that the ALP-2 exerted these inhibitory effects at an early phase of B cell proliferation and differentiation. Nevertheless, the proliferative response of thymocytes to rIL-4 was not inhibited by ALP-2. In addition, ALP-2 inhibited neither the B cell proliferation induced by interleukin 2 nor the B cell differentiation by interleukin 5. Cross-inhibition experiments, together with immunoblotting analysis, revealed that the LP-2 recognized by ALP-2 appears to be an epitope of CD45R (B220) molecule(s), the isoform(s) of the Ly-5 antigen system. Epitope mapping analysis using CD45 gene transfectants showed that Lp-2 epitope is dependent upon the expression of the first alternatively spliced exon of CD45 gene. Functional studies showed that the mAb to an epitope of CD45R, RA3-2C2, but not RA3-6B2, had similar inhibitory effects on the IL-4-mediated proliferative response of B cells. These findings suggest that the CD45R molecules associated with Lp-2 and RA3-2C2 epitopes are probably related to a signal transduction provided by IL-4 in B cells. The possibility that different pathways are operative for IL-4-mediated signaling between B and T cells has to be considered.     

Monitoring of antigen-specific CD8 T cells in patients with type 1 diabetes treated with antiCD3 monoclonal antibodies

The way in which anti-CD3 monoclonal antibodies (mAbs) modify human immune responses in type 1 diabetes (T1DM) is not known. We prepared a panel of Class I HLA-A2.1 tetramers with peptides from diabetes-associated antigens and studied the frequency and phenotype of the cells in patients with T1DM and blood donors and in patients treated with anti-CD3 mAb (Teplizumab). More patients with T1DM showed positive staining for at least 1 tetramer using frozen and fresh samples (p < 0.05). Three months following treatment with anti-CD3 mAb, the proportion of GAD65- and InsB-peptide reactive CD8+ T cells increased (p < 0.05). The phenotype of these cells was modulated from naïve to effector memoryRA+. We concludethat Class I MHC tetramers can identify antigen specific CD8+ T cells in patients with T1DM. The frequency of certain specificities increases after treatment with anti-CD3 mAb. Their modulated phenotype may have functional consequences for their pathogenicity.     

抗B7-1和抗CD40L单抗延长小鼠皮肤移植物存活实验研究

将C57BL/6小鼠腹部全层皮肤移植于BALB/c小鼠中段背部,实验分组(每组7只小鼠):1.B7-1功能性单抗(4E5)治疗组;2.抗CD40L单抗(4F1)治疗组;3.4E5+4F1治疗组,各单抗以20㎎/㎏的剂量注入腹腔内;4.空白对照组,只注射同等量的生理盐水。注射时间为移植后0﹑1﹑3﹑5d,观察移植皮肤排斥情况。于术后第6天分别杀死各组受体和供体鼠,取受体脾细胞与供体作混合淋巴细胞反应(MLR)。收集培养6d的初次反应细胞,检测再次MLR。结果发现,与对照组相比,各单抗治疗组皮肤移植物存活时间延长(P<0.05﹚;与各单独应用4F1和4E5相比,联合使用4F1和4E5产生一定的协同作用,但未能进一步延长移植物的寿命(P>0.05)。初次单向MLR:4F1﹑4E5和4F1+4E5治疗组受体T淋巴细胞在MLR中表现对供体淋巴细胞特异性低反应性,能有效抑制T细胞对同种异体抗原的初次应答。再次单向MLR:4F1﹑4E5﹑4F1+4E5对供体淋巴细胞在再次反应中仍保持着对同种抗原的反应性,与对照组无显著差异,未能诱导特异性免疫耐受。综上结果证实,anti-CDB7-1mAb(4E5)和anti-CD40LmAb(4F1)作为新型免疫抑制剂,在一定程度上抑制细胞免疫应答,干预排斥反应。     

PD-L1 expression analysis in gastric carcinoma tissue and blocking of tumor-associated PD-L1 signaling by two functional monoclonal antibodies

Programmed death-1 ligand-1 (PD-L1), a member of the B7 family of costimulatory molecules, plays an important role in the regulations of the cellular and humoral immune responses. In this study, two mouse anti-human PD-L1 monoclonal antibodies named 10E10 and 2H11 were successfully generated and further characterized. Monoclonal antibody 10E10 bound to distinct PD-L1 epitope comparing an available anti-PD-L1 monoclonal antibody on a series of malignant cell lines, activated T lymphocytes, B lymphocytes and dendritic cells. Then, by using immunohistochemistry staining with monoclonal antibody 2H11, the expression of PD-L1 was found in human gastric carcinoma specimens but not in normal or gastric adenoma tissues. Additional data show that PD-L1 can be regarded as a decisive factor in evaluating gastric carcinoma prognosis and anti-human PD-L1 monoclonal antibody 10E10 could inhibit T-cell apoptosis induced by tumor-associated PD-L1. Taken together, these results showed that the two functional mouse anti-human PD-L1 monoclonal antibodies we generated might be of great value for further exploration of the costimulatory molecule regulating network and immunointervention for tumor immunotherapy.     

A triple antibody assay for the quantitation of plasma IgG subclass antibodies to acetylcholine receptors in patients with myasthenia gravis

Monoclonal anti-human IgG subclass antibodies have been used in an immunoprecipitation assay for the determination of anti-acetylcholine receptor IgG subclasses in plasma from patients with myasthenia gravis. Solubilized acetylcholine receptors labelled with 125I-alpha-bungarotoxin were incubated with patient plasma. Monoclonal mouse antibodies to human IgG subclasses 1-4 were added to the incubation and finally precipitated with anti-mouse IgG antibody. A maximal IgG subclass precipitation of 62-76% was determined with 125I-labelled myeloma IgG subclasses 1-4 added to normal human plasma. The anti-IgG subclass antibodies were added in excess which ensured that the precipitation of IgG2, IgG3 or IgG4 were unchanged, and that of IgG1 was only reduced by 17%, when the plasma IgG concentration was increased by a factor of two. The anti-IgG subclass antibodies were highly specific for their complementary subclasses. Determination of the IgG subclass of the anti-acetylcholine receptor antibodies from 8 patients with myasthenia gravis showed that IgG1 and IgG3 antibodies are predominant. This may support the hypothesis that complement mediated lysis of the neuromuscular end-plate plays a pathogenetic role in myasthenia gravis.     

Immunohistochemical detection of cd1a antigen in formalin-fixed and paraffin-embedded tissue sections with monoclonal antibody 010

The immunoreactivity of a CD1a monoclonal antibody (MAb), denoted 010, was investigated by means of the streptavidin-biotin-peroxidase method in formalin-fixed and paraffin-embedded tissues from 47 cases. The samples comprised reactive lymphoid proliferations of skin, tonsil, and lymph node including dermatopathic lymphadenopathy and Langerhans' cell histiocytosis, Hodgkin's and non-Hodgkin's lymphomas, and thymomas. Interdigitating and dermal dendritic cells, veiled cells, Langerhans' cells, and also cortical thymocytes and their neoplastic counterparts displayed immunostaining with MAb 010 in paraffin sections. These results are identical to previous ones reported for other CD1a MAbs in fresh or frozen specimens. The findings suggest that the binding site of 010 is a fixation-resistant epitope of CD1a antigen which has not been previously identified.     

Immunohistochemical results of HER2/neu protein expression assessed by rabbit monoclonal antibodies SP3 and 4B5 in colorectal carcinomas

HER2/neu is an efficient target for cancer therapy. However, reports about its overexpression rate in colorectal carcinomas showed wide variability. This study aims to investigate HER2/neu expression in colorectal carcinomas using these two rabbit monoclonal HER2/neu antibodies, and to clarify the relationship between protein overexpression and gene amplification of HER2/neu and their clinicopathologic importance. Tissue microarray was performed from sections of 106 cases colorectal carcinomas. Their clinical data, including gender, age, stage, recurrence, lymph node metastasis, and follow-ups were collected. Immunohistochemistry for rabbit monoclonal antibody SP3 and 4B5 were performed, Fluorescent in situ hybridization was applied to detect the amplification of HER2/neu gene. The HER2/neu overexpression of (2+ and 3+) in our results were seen in 7.5% (8/106) for 4B5 and 3.8% (4/106) for SP3 respectively, the HER2/neu amplification was in 2.8% (3/106). All cases of overexpression for SP3 were included by those for 4B5. Both antibodies stained 3 cases of HER2/neu 3+, and FISH confirmed HER2/neu amplification did occurred in these cases. In our study, 4B5 was more sensitive to detect HER2/neu of colorectal carcinoma than SP3. 2.8% patients with colorectal patients might benefit from anti-HER2/neu therapy.