Anti-Kinesin Heavy Chain antibody [KN-01] (ab9097)

Immunocytochemistry/ Immunofluorescence - Kinesin Heavy Chain antibody [KN-01] (ab9097)

This antibody gives typical dot-like staining characteristic for localization of kinesin in vesicles. Optimum staining of cell lines was observed on samples fixed with formaldehyde followed by Triton X-100 extraction.

Immunofluorescence staining of embryonal mouse fibroblasts 3T3 with MAb KN-01. Double-label fluorescence with polyclonal affinity-purified anti-tubulin antibody reveals a co-distribution of vesicles with microtubules on the cell periphery. Vesicles were stained with anti-Kinesin Heavy Chain MAb KN-01 (red), microtubules with anti-tubulin Ab (green) and nucleus with DAPI (blue).

Dr. E. Dráberová, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Kinesin Heavy Chain antibody [KN-01] (ab9097)

ab9097 (4µg/ml) staining Kinesin Heavy Chain in human testis using an automated system (DAKO Autostainer Plus). Using this protocol there is very weak cytoplasmic staining seminal vesicle.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

Flow Cytometry-Anti-Kinesin Heavy Chain antibody [KN-01](ab9097)

Overlay histogram showing HeLa cells stained with ab9097 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab90970, 1µg/1x10⁶ cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgM [ICIGM] (ab91545, 2µg/1x10⁶ cells ) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.