Anti-Tubulin抗体[YOL1/34] - Microtubule Marker (ab6161)

ab6161 staining Tubulin in HeLa cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab6161 at 5µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150165, Goat polyclonal Secondary Antibody to Rat IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor^® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

Western blot against tubulin with ab6161 at 1/3000.  Secondary Rabbit anti-Rat IgG HRP (ab6734)was used at 1/2000.  Exposure time: 2mins.

Lane 1: 20µg/lane HeLa (Human) whole cell lysates (ab7898).

Lane 2: 20µg/lane 3T3 (Mouse) whole cell lysate (ab7901).

Lane 3: 20µg/lane Rat brain tissue lysate (ab7942).

Confocal image of  21 day in vitro rat hippocampal neurons, stained with rat monoclonal antibody to Tubulin - Microtubule Marker (ab6161) in green at 1/500 and Microtubule Associated protein 2 in blue.

This picture was kindly supplied as part of the review submitted by Dr Jonathon Burman.

Cultured human macrophages were used with ab6161 at 1/1000 for immunofluorescence. Cells were fixed with cold 2% formaldehyde for 20mins.

Green staining is Alexa 568, Blue staining is DAPI stain.

This cell represents a young macrophage, the staining patterns varied as the cells aged in culture.

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ICC/IF image of ab6161 stained human HepG2 cells. The cells were 4% PFA fixed (10 min), permabilised in 0.1% PBS-Tween (20 min) and incubated with the antibody (ab6161, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor^® 488 goat anti-rat IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor^® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue). This antibody also gave a positive IF result in HeLa, HEK 293 and MCF7 cells.

ab6161 staining mouse NIH 3T3 fibroblast cells by ICC/IF. Cells were PFA fixed  and permeabilized in 0.2% Triton X-100 prior to blocking in 5% BSA for 45 minutes at RT.  The primary antibody was diluted 1/1000 and incubated with the sample for 1 hour.  An Alexa Fluor^® 568 conjugated goat anti-rat antibody, diluted 1/3000, was used as the secondary.

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