Published customer image: Mouse anti Human CD49d antibody, clone HP2/1 used for western blotting and immunoprecipitationImage caption:Tetraspanins CD81, CD82 and CD151 are associated with a4beta1 throughout erythroid maturation and with beta3 in proerythroblasts and basophilic erythroblasts. A. CD81, CD82 and CD151 precipitates from Mn2+-activated proerythroblasts (ProEB, day 5), basophilic (BasoEB, day 8) and polychromatic (PolyEB, day 12) erythroblasts were successively probed with anti-a4, anti-beta1 and anti-beta3 antibodies; tetraspanin controls from each time point are also illustrated. All tetraspanins co-precipitated a4 and beta1 from erythroblasts B. Tetraspanin precipitates from day 6 proerythroblasts (ProEB) solubilised in the presence of EDTA or different cations, and from Mn2+-activated basophilic erythroblasts (BasoEB, day 8) were probed with a mix of antibodies to a5, beta1, beta2 and beta3 integrins while the control samples were probed with the relevant tetraspanin antibodies. For clarity, integrin controls are illustrated for the EDTA blot but were present on all blots. beta1 and beta3 integrins were precipitated well only in the presence of Mn2+. C. CD81 and CD82 precipitates from day 5 proerythroblasts were successively probed with different anti-integrin subunit antibodies and demonstrate co-precipitation of beta1 and beta3 but not a5 or beta2 integrins. D. CD81 (454720) and CD82 (53H5) precipitates from day 6 proerythroblasts (ProEB) and HEL cells (HEL) solubilised in the presence of EDTA, Ca2++Mg2+ or Mn2+ probed with anti-CD82 and anti-CD81 antibodies. Each tetraspanin co-precipitates the other most strongly in the presence of Mn2+ from proerythroblasts while any cation permits co-precipitation in HEL cells. Integrins were analysed on 7.5% gels, tetraspanins on 12% gels; non-reducing conditions. Unless stated, the following clones were used: CD53, MEM-53; CD63, MEM-259; CD81, 454720; CD82, TS82b; CD151, IIG5a; a4, HP2/1; a5, IIA1; aL, TS1/22; aIIb, PAB-1. All day 5 and 6 cultures comprised 90 -95% proerythroblasts; day 8 culture comprised 5% proerythroblasts, 81% basophilic erythroblasts and 14% polychromatic erythroblasts; day 12 culture comprised 41% polychromatic erythroblasts, 15% orthochromatic erythroblasts and 41% reticulocytes. In the day 5 and 6 cultures 15 -34% of cells were GPA+ and 28 -35% of cells were aIIb+. Day 8 and day 12 cultures had 77% and 97% GPA+ cells, respectively, and 9% and 0% aIIb+ cells, respectively.From: Spring FA, Griffiths RE, Mankelow TJ, Agnew C, Parsons SF, et al. (2013) Tetraspanins CD81 and CD82 Facilitate a4beta1-Mediated Adhesion of Human Erythroblasts to Vascular Cell Adhesion Molecule-1. PLoS ONE 8(5): e62654.

Published customer image: Mouse anti Human CD49d antibody, clone HP2/1 used for immunoprecipitationImage caption:All anti-CD81, anti-CD82 and anti-CD151 clones co-precipitate beta1 and beta3 integrins from normal and leukemic proerythroblasts. A. CD81 clones. B. CD82 clones. C. CD151 clones. ERB, day 6 proerythroblasts. a4, HP2/1; a5, IIA1; aL, TS1/22; aIIb, PAB-1. Integrins in 7.5% NR gels, tetraspanins in 12% non-reduced gels. More beta3 integrins are co-precipitated from HEL cells as they express aIIbbeta3 and aVbeta3; proerythroblasts express only aIIbbeta3.From: Spring FA, Griffiths RE, Mankelow TJ, Agnew C, Parsons SF, et al. (2013) Tetraspanins CD81 and CD82 Facilitate a4beta1-Mediated Adhesion of Human Erythroblasts to Vascular Cell Adhesion Molecule-1. PLoS ONE 8(5): e62654.

Published customer image: Mouse anti Human CD49d antibody, clone HP2/1 used for immunoprecipitationImage caption:All anti-CD81, anti-CD82 and anti-CD151 clones co-precipitate beta1 and beta3 integrins from normal and leukemic proerythroblasts. A. CD81 clones. B. CD82 clones. C. CD151 clones. ERB, day 6 proerythroblasts. a4, HP2/1; a5, IIA1; aL, TS1/22; aIIb, PAB-1. Integrins in 7.5% NR gels, tetraspanins in 12% non-reduced gels. More beta3 integrins are co-precipitated from HEL cells as they express aIIbbeta3 and aVbeta3; proerythroblasts express only aIIbbeta3.From: Spring FA, Griffiths RE, Mankelow TJ, Agnew C, Parsons SF, et al. (2013) Tetraspanins CD81 and CD82 Facilitate a4beta1-Mediated Adhesion of Human Erythroblasts to Vascular Cell Adhesion Molecule-1. PLoS ONE 8(5): e62654.

Published customer image: Mouse anti Human CD49d antibody, clone HP2/1 used for blocking studies.Image caption: Blocking experiments with anti-beta7, anti-a4 and anti-aVbeta3 antibodies for adhesion to HS-5 stromal cells. Panel A, Blocking with anti-beta7. HMCLs were stimulated with Pam3CSK4 for 24 hours and then treated with anti-beta7 antibody before adhesion to HS5-coated wells. Panel B, Blocking experiments with anti-a4 and anti-aVbeta3 antibodies for adhesion to HS-5. HMCLs were stimulated with Pam3CSK4 for 24 hours and then treated with anti-a4 and anti-aVbeta3 antibodies before adhesion to HS5-coated wells. The results are the statistical analyses of data in 3 separate experiments expressed as mean +/- SEM, *P

Published customer image: Mouse anti Human CD49d antibody, clone HP2/1 used for binding efficiency determinationImage caption:Binding efficiencies (BE) of different a4beta7 molecules composed of distinct a4 mutants to monoclonal antibodies against a4, beta7 or the a4beta7 heterodimer. Binding efficiency is determined by the ratio between the mean fluorescence of antibody binding to each a4 molecule and of the binding in a mock-transfected cell culture (see Materials and Methods for details). Dark gray bars represent binding to the human (wild type) a4 clone, whereas light gray bars are those of binding to the different a4 mutants (as shown in the x-axis). a4 mutants which included substitutions at codon 201 are boxed. A, binding of anti-a4 2b4 antibody. B, binding of anti-a4 HP2/1 antibody. C, BE of different anti-a4 and beta7 antibodies to the human a4 and the quintuple a4 mutant (5 aa mut). Bars represent the range of standard errors deduced from triplicate experiments. p-values of Student's t tests are shown above each comparison. NS, non-significant (> 0.05).From:Darc M, Hait SH, Soares EA, Cicala C, Seuanez HN, et al. (2011) Polymorphisms in the a4 Integrin of Neotropical Primates: Insights for Binding of Natural Ligands and HIV-1 gp120 to the Human a4beta7. PLoS ONE 6(9): e24461.