<b>OBJECTIVEb>To investigate the effects of T cell receptor (TCR) BD2-BJ2 gene rearrangement on the complementary-determining region (CDR) 3 of TCR beta chain (TCR BV CDR3) in Jurkat cells.

<b>METHODSb>TCR BV gene subfamilies were detected by RT-PCR in Jurkat cells during proliferation and after induction with non-specific T cell activators and SEA, respectively. To determine the clonality of TCR BV subfamilies and the lengths of CDR3, the PCR products were analyzed by TCR GeneScan technique, and the sequences of CDR3 were further analyzed by DNA sequencer.

<b>RESULTSb>No new TCR BV subfamilies were found in Jurkat cells, a monoclonal BV8(+)cell line, either during cell proliferation or after stimulation with different treatments, nor were any differences found in CDR3 size or sequences.

<b>CONCLUSIONb>TCR BD2-BJ2 rearrangement in Jurkat cells may not play a role in modification of TCR BV CDR3 domains or the consequent antigen immunorecognition of BV CDR3, but the possibility of TCR modification can not be excluded.

Amino Acid Sequence ; Base Sequence ; Complementarity Determining Regions ; genetics ; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor ; genetics ; Humans ; Immunologic Factors ; genetics ; Jurkat Cells ; Leukemia, T-Cell ; genetics ; immunology ; pathology ; Molecular Sequence Data ; Receptors, Antigen, T-Cell ; genetics ; T-Lymphocytes ; immunology ; metabolism

<b>UNLABELLEDb>The purpose of this study was to evaluate the effects of cellular immunity activation on P58(+) cells expressing killer cell inhibitory receptor (KIR) and their regulatory function on cellular immunity, and provid theoretical data for preventing graft-vers-host disease (GVHD) in stem cell transplantation therapy. The mononuclear cells from human peripheral blood were incubated with IL-2, Con A and Lipostin (LP) for 72 hours. The KIR expressing cells, P58.1(+) and P58.2(+) cells, were analyzed by flow cytometry. The results showed that the percentages of CD3(+), CD4(+), CD8(+), CD16(+)CD56(+), P58.1(+) and P58.2(+) cells were greatly increased after treated with IL-2, Con A and LP, separately or in combination, and the percentages of above cells in combined treatment groups were higher than those of single stimulated groups, especially the percentage of cells in the IL-2 + LP group was significantly higher than those in IL-2 and LP singly treated groups.

<b>IN CONCLUSIONb>IL-2, Con A and LP possess the ability to induce the expression of KIR and stimulate proliferation of P58.1(+) and P58.2(+) cells while to activate the celluar immunity response, the expression of P58 gene may be regulated by the activation of cellular immunity.

Adult ; CD3 Complex ; analysis ; CD4 Antigens ; analysis ; CD56 Antigen ; analysis ; CD8 Antigens ; analysis ; Cell Count ; Cell Division ; drug effects ; Concanavalin A ; pharmacology ; Flow Cytometry ; Humans ; Interleukin-2 ; pharmacology ; Leukocytes, Mononuclear ; cytology ; drug effects ; immunology ; Receptors, IgG ; analysis ; Receptors, Immunologic ; analysis ; Receptors, KIR ; Receptors, KIR2DL3

<b>OBJECTIVEb>To analyze the drift of the complementarity-determining region 3 (CDR3) of T cell receptor beta (TCRbeta) chain variable region in T cells of healthy volunteers cultured with interleukin-2 (IL-2).

<b>METHODSb>T cells were isolated from the peripheral blood and cultured in vitro in the presence of IL-2. The non-specific killing effect of the cells was analyzed by LDH releasing assay, and the distribution of TCRbeta chain CDR3 in healthy volunteers by immunoscope spectratyping method to evaluate the clonality of the T cells.

<b>RESULTSb>The results showed Gaussian distribution of TCR Vbeta gene CDR3 in healthy volunteers. The T cell cultured with IL-2, however, displayed some anomalous and oligoclonal expansion in different TCR Vbeta families without killing effect against nasophargngal carcinoma cell line CNE2.

<b>CONCLUSIONb>IL-2 may affect TCRbeta chain CDR3 distribution in T cells cultured in vitro.

Cells, Cultured ; Complementarity Determining Regions ; genetics ; Genetic Drift ; Humans ; Interleukin-2 ; metabolism ; Leukocytes, Mononuclear ; metabolism ; Receptors, Antigen, T-Cell, alpha-beta ; genetics ; T-Lymphocytes ; immunology ; metabolism

<b>OBJECTIVEb>To explore the most efficient culture system which can induce cord blood (CB)-mononuclear cells (MNC) to differentiate into mature T/NK cells in vitro.

<b>METHODSb>The CB MNCs were cultured in six culture systems respectively for 4 weeks. The T/NK cell surface phenotypes were analyzed by flow cytometry and the absolute numbers of nucleated cells (NCs) were counted at each time point. Moreover, cell morphology was identified by Giemsa-Wright staining, and cytotoxicity of the cultured cells to K562 and Raji tumor cells was also evaluated by MTT method.

<b>RESULTSb>Cultured in the cytokine cocktail of SCF + FLT-3L + IL-7 + IL-15 + TNF-alpha + IL-2, the NCs were (20 approximately 26) x 10(6)/ml in numbers at day 22. The percentage of lymphocytes in the NCs and that of CD(3)(+) T cells in the lymphocytes both exceeded 90% at the same time. Most of the CD(3)(+) T cells were CD(3)(+)CD(8)(+) and the percentage of CD(3)(+)CD(4)(+) T cells declined gradually. The percentage of CD(3)(+)CD(56)(+) NKT cells and gamma delta(+)T cells in the lymphocytes arised from lower than 2% to 30% approximately 40% and 10% approximately 15%, respectively. CD(3)(-)CD(56)(+) NK cells were not expanded. The cytotoxic activity of the cultured cells to K562 and Raji cells at an effector:target (E:T) ratio of 50:1 was over 75% and about 32% approximately 65%, respectively.

<b>CONCLUSIONb>The most efficient culture system which can induce CB MNC to differentiate into mature T/NK cells in vitro is the cytokines cocktail of SCF + FLT-3L + IL-7 + IL-15 + TNF-alpha + IL-2, and the optimum culture time is 22 days.

Cell Differentiation ; Cell Division ; Cytotoxicity, Immunologic ; Fetal Blood ; cytology ; Humans ; Infant, Newborn ; Interleukin-2 ; pharmacology ; Killer Cells, Natural ; cytology ; Leukocytes, Mononuclear ; cytology ; Receptors, Antigen, T-Cell, gamma-delta ; analysis ; T-Lymphocytes ; cytology ; Tumor Necrosis Factor-alpha ; pharmacology

<b>OBJECTIVESb>To understand the role cellular immunology plays in the pathogenesis of chronic hepatitis B (CHB) through analysis of T cell receptor (TCR) beta chain variable region gene (BV) family dominant usage and beta chain complementarity determining region3 (CDR3) sequences of peripheral blood mononuclear cells of the patients.

<b>METHODSb>TCR BV families were amplified by inverse polymerase chain reaction (RT-PCR), and the dominant usage of BV families and CDR3 repertoire were analyzed by immunoscope technology for 8 CHB patients during their acute exacerbations and for 4 healthy blood donors who served as controls. The clonality of the T cells suspected by immunoscope was further confirmed by CDR3 sequencing.

<b>RESULTSb>The TCR BV CDR3 repertoire of the 4 healthy blood donors showed a Gaussian distribution. In the 8 CHB patients, however, the clonal expansion of T cells showed different TCR BV families with each patient. The T cells of the clonal expansion shared different CDR3 sequences.

<b>CONCLUSIONb>The peripheral blood T cells of CHB patients during their acute exacerbation showed significantly a clonal expansion and their T cell clonal expansion may be stimulated by several HBV epitopes. These results indicate that cellular immunology is involved in the pathogenesis of the liver inflammation process of CHB.

Adult ; Amino Acid Sequence ; Base Sequence ; Cloning, Molecular ; Complementarity Determining Regions ; genetics ; Female ; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor ; genetics ; Hepatitis B, Chronic ; genetics ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; Receptors, Antigen, T-Cell ; genetics

<b>OBJECTIVEb>To analyze the characteristics of CDR3 of TCRbeta on CD8+ T cells in chronic hepatitis B patients.

<b>METHODSb>Eight patients with chronic hepatitis B (ALT more than 2 ULN) were enrolled in this study. CD8+ T cells were isolated from peripheral blood. RT-PCR was proformed to amplify the CDR3 of TCRbeta, and the PCR products were sequenced and analyzed.

<b>RESULTSb>The chronic hepatitis B patients showed obvious clonal expansion of T cell, and three perturbation patterns of T cell expansion were showed in the CDR3 of TCRbeta, including monoclonicity, oligoclonicity and skewed peak patterns. The number of perturbation families of CD8+ subpopulation was significantly higher than that of CD8- subpopulation (10.6+/-4.7 vs. 4.1+/-3.1, t = 6.619, P less than 0.01). In 3 out of 8 patients, the number of perturbation families of CD8+ subpopulation was also higher than that of PBMCs without depleting CD8+ subpopulation.

<b>CONCLUSIONSb>The characteristics of CDR3 of TCRbeta may help to understand the inflammatory response in CHB patients.

Adult ; CD8-Positive T-Lymphocytes ; immunology ; Complementarity Determining Regions ; genetics ; Genes, T-Cell Receptor beta ; Hepatitis B, Chronic ; blood ; immunology ; Humans ; Male ; Receptors, Antigen, T-Cell ; immunology ; Young Adult

<b>OBJECTIVEb>To study the expression of TCR BV CDR3 family in fulminant hepatitis B (FHB) patients.

<b>METHODSb>Totally 28 patients with fulminant hepatitis B (FHB) (FHB group), who were treated in our hospital from Oct. 2010. to Mar. 2012, and 20 healthy controls( HC group) were included in the study. PBMCs were isolated from anticogulated blood, and RT-PCR was used to detect the levels of TCR BV CDR3 family in the 2 groups.

<b>RESULTSb>The levels of DeltaCt1, DeltaCt12 and DeltaCt20 in FHB group were higher than those in HC group (P < 0.05); The levels of DeltaCt5, DeltaCt7, DeltaCt13, DeltaCt14, DeltaCt15, DeltaCt22, DeltaCt23 in FHB group were lower than those in HC group (P < 0.05).

<b>CONCLUSIONSb>The result indicates that cellular immunology is involved in the pathogenesis of the liver inflammation process of FHB.

Adolescent ; Adult ; Aged ; Case-Control Studies ; Complementarity Determining Regions ; chemistry ; genetics ; immunology ; Female ; Hepatitis B ; genetics ; immunology ; virology ; Humans ; Male ; Middle Aged ; Receptors, Antigen, T-Cell ; chemistry ; genetics ; immunology ; Young Adult

There are many factors that can influence the pharmacokinetics (PK) of a mAb or Fc-fusion molecule with the primary determinant being FcRn-mediated recycling. Through Fab or Fc engineering, IgG-FcRn interaction can be used to generate a variety of therapeutic antibodies with significantly enhanced half-life or ability to remove unwanted antigen from circulation. Glycosylation of a mAb or Fc-fusion protein can have a significant impact on the PK of these molecules. mAb charge can be important and variants with pI values of 1-2 unit difference are likely to impact PK with lower pI values being favorable for a longer half-life. Most mAbs display target mediated drug disposition (TMDD), which can have significant consequences on the study designs of preclinical and clinical studies. The PK of mAb can also be influenced by anti-drug antibody (ADA) response and off-target binding, which require careful consideration during the discovery stage. mAbs are primarily absorbed through the lymphatics via convection and can be conveniently administered by the subcutaneous (sc) route in large doses/volumes with co-formulation of hyaluronidase. The human PK of a mAb can be reasonably estimated using cynomolgus monkey data and allometric scaling methods.
Absorption, Physiological ; Animals ; Antibodies, Monoclonal ; pharmacokinetics ; Dose-Response Relationship, Immunologic ; Humans ; Receptors, Fc ; metabolism ; Recombinant Fusion Proteins ; pharmacokinetics ; Tissue Distribution

To rapidly select potent anti-VSTM1-v2 scFv (single-chain antibody fragment) by construction and screening of a humanized scFv library in which a murine VH-CDR3 library was grafted onto a human scFv framework. A murine VH-CDR3 library was amplified from anti-VSTM1-v2 murine cDNA and grafted on human scFv (VH3-VK1) framework. Anti-VSTM1-v2 scFv templates were selected and enriched through ribosome display, TA-cloned into expression vector, and transformed into BL21 (DE3) for soluble expression of target scFv. A total of 1000 clones were randomly picked. Positive ones were first identified using colony PCR, indirect ELISA, Western blotting and then verified with sequencing and dose response ELISA. At last an anti-VSTM1-v2 humanized scFv with good binding affinity (EC50 = 21.35 nmol x L(-1)) was selected from the humanized library of 10(12) members generated in this study. This scFv antibody might have potential applications. This study provides a new approach for rapid screening of humanized antibodies.
Animals ; Complementarity Determining Regions ; genetics ; immunology ; Cytokines ; immunology ; Humans ; Immunoglobulin Fragments ; genetics ; immunology ; Immunoglobulin Heavy Chains ; genetics ; immunology ; Mice ; Peptide Library ; Protein Binding ; Receptors, Immunologic ; immunology ; Single-Chain Antibodies ; genetics ; immunology ; isolation & purification

B cell activated co-receptor plays important roles in linkage of innate and acquired humoral immune responses. CD21 molecule in the co-receptor complex is a receptor for C3dg and CD19 molecule enhances BCR signal transduction. CD21 also expresses on the surface of follicular dendritic cells, which mediates the long-term maintenance of antigens and is indispensable for maintaining the memory of B cells. B cell activated co-receptor also has an effect on the negative selection of B cells reactive to autoantigens.
Animals ; Antigens, CD19 ; immunology ; Autoantigens ; immunology ; B-Lymphocytes ; immunology ; Humans ; Receptors, Complement 3d ; immunology ; Receptors, Immunologic