Immunocytochemistry/ Immunofluorescence - HA tag antibody - ChIP Grade (ab9110)

This image was kindly supplied as part of the review submitted by Kasper Fugger. Immunofluorescence staining of U-2 cells expressing HA-tagged protein with ab9110.

ChIP - HA tag antibody - ChIP Grade (ab9110)

Xenopus laevis oocytes were injected with mRNA for HA-tagged human BORIS. Chromatin was prepared according to the Abcam X-ChIP protocol. Oocytes were fixed with formaldehyde for 10min. The  ChIP was performed with 25µg of chromatin, 20µl of Protein A/G sepharose beads, and 3µg of ab9110 (anti-HA, light blue) or, 3µg of ab18337 (anti-Boris, dark blue). A non-specific antibody was used as a control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach).

Immunocytochemistry/ Immunofluorescence - HA tag antibody - ChIP Grade (ab9110)

ab 9110 at a 1/200 dilution staining the mouse olineu cell line (oligodendrocyte precursor cell) by immunocytochemistry. The antibody was incubated with the cells for 30 minutes and then detected using a Cy5 conjugated goat anti-rabbit antibody.

This image is courtesy of an Abreview submitted by Katarina Trajkovic on 15 March 2006

Western blot - HA tag antibody - ChIP Grade (ab9110)

All lanes : Anti-HA tag antibody - ChIP Grade (ab9110) at 1/4000 dilution

Lane 1 : 15ug untransfected wcl lysate
Lane 2 : 293FT cells transfected with 15kDa HA tagged Vpr (an HIV1 accessory protein)

Secondary
HRP conjugated Goat anti-Rabbit
developed using the ECL technique

Performed under reducing conditions.

Observed band size : 15 kDa (why is the actual band size different from the predicted?)


Exposure time : 5 seconds

This image is courtesy of an Abreview

Immunoprecipitation - HA tag antibody - ChIP Grade (ab9110)

ab9110 was diluted to 4 µg/mg lysate and incubated with a nuclear lysate of HEK293T cells transiently expressing HA-tagged protein and a Protein A matrix for 2 hours a 23°C to achieve immunoprecipitation.  1000 µg of lysate was present in the input. 

A HRP-conjugated anti-rabbit HA monoclonal antibody diluted 1/1000 was used for the Western Blot step.

This image is courtesy of an anonymous Abreview