TTC34 Antibody / Tetratricopeptide repeat protein 34 (FY12119)

Immunofluorescent staining of TTC34 using anti-TTC34 antibody (green) and anti-Beta Tubulin antibody (red). TTC34 was detected in an immunocytochemical section of U2OS cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 ug/ml rabbit anti-TTC34 antibody and mouse anti-Beta Tubulin antibody overnight at 4oC. DyLight 488 Conjugated Goat Anti-Rabbit IgG and DyLight 594 Conjugated Goat Anti-Mouse IgG were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37oC. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Western blot analysis of TTC34 using anti-TTC34 antibody. Lane 1: human Jurkat whole cell lysates, Lane 2: rat testis tissue lysates, Lane 3: rat PC-12 whole cell lysates, Lane 4: rat C6 whole cell lysates, Lane 5: mouse testis tissue lysates, Lane 6: mouse NIH/3T3 whole cell lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-TTC34 antibody at 0.5 ug/ml overnight at 4oC, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal was developed using enhanced chemiluminescent. The expected band size for TTC34 is at 61 kDa but it is commonly observed at 48-55 kDa due the proteins TPR-repeat/highly acidic/low SDS-binding regions composition causing it to migrate faster in SDS-PAGE.

IHC analysis of TTC34 using anti-TTC34 antibody. TTC34 was detected in a paraffin-embedded section of human cervical cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 ug/ml rabbit anti-TTC34 antibody overnight at 4oC. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37oC. The tissue section was developed using an HRP secondary and DAB substrate.

Flow Cytometry analysis of Jurkat cells using anti-TTC34 antibody. Overlay histogram showing Jurkat cells stained with (Blue line). The cells were fixed with 4% paraformaldehyde and blocked with 10% normal goat serum. And then incubated with rabbit anti-TTC34 antibody (1 ug/million cells) for 30 min at 20oC. DyLight 488 conjugated goat anti-rabbit IgG (5-10 ug/million cells) was used as secondary antibody for 30 minutes at 20oC. Isotype control antibody (Green line) was rabbit IgG (1 ug/million cells) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.