c-Fos Recombinant Rabbit Monoclonal Antibody [PSH07-51]

Application: IHC-Fr

Species: Mouse

Site: Hypothalamus (restraint stress induced)

Sample: Frozen section

Antibody concentration: 1/4,000

Antigen retrieval: Not required

Important Notice: Blocking buffer and antibody dilution buffer are recommended to use TBS buffer instead of PBS buffer.

Application: IHC-Fr

Species: Mouse

Site: Cerebral cortex (restraint stress induced)

Sample: Frozen section

Antibody concentration: 1/4,000

Antigen retrieval: Not required

Important Notice: Blocking buffer and antibody dilution buffer are recommended to use TBS buffer instead of PBS buffer.

Application: IHC-Fr

Species: Rat

Site: Cerebral cortex (restraint stress induced)

Sample: Frozen section

Antibody concentration: 1/4,000

Antigen retrieval: Not required

Important Notice: Blocking buffer and antibody dilution buffer are recommended to use TBS buffer instead of PBS buffer.

Western blot analysis of c-Fos on different lysates with Rabbit anti-c-Fos antibody (HA722839) at 1/5,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: HeLa serum starved for 16 hours then add 200 nM TPA for 4 hours cell lysate
Lane 3: RAW264.7 cell lysate
Lane 4: RAW264.7 serum starved for 16 hours then add 200 nM TPA for 4 hours cell lysate

Lysates/proteins at 15 µg/Lane.

Predicted band size: 41 kDa
Observed band size: 41-55 kDa

Exposure time: 10 seconds; ECL: K1801;

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722839) at 1/5,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

Immunohistochemical analysis of paraffin-embedded mouse brain (restraint stress induced) tissue with Rabbit anti-c-Fos antibody (HA722839) at 1/5,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722839) at 1/5,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Immunohistochemical analysis of paraffin-embedded rat brain (restraint stress induced) tissue with Rabbit anti-c-Fos antibody (HA722839) at 1/5,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722839) at 1/5,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Immunocytochemistry analysis of HeLa cells serum starved for 40 hours then add 20% FBS for 2 hours labeling c-Fos with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Immunocytochemistry analysis of RAW264.7 cells serum starved for 16 hours then add 20% FBS for 4 hours labeling c-Fos with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Immunocytochemistry analysis of C6 cells serum starved for 16 hours then add 10% FBS for 30 minutes labeling c-Fos with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-c-Fos antibody (HA722839) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells (serum starved for 16 hours and treated with 200 nM TPA for 4 hours) with c-Fos (HA722839) or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

Application: IF-tissue

Species: Mouse

Site: Cerebral cortex (restraint stress induced)

Sample: Paraffin-embedded section

Antibody concentration: 1/4,000

Important Notice: Blocking buffer and antibody dilution buffer are recommended to use TBS buffer instead of PBS buffer.

c-Fos was immunoprecipitated from 0.2 mg HeLa serum starved for 40 hours then add 20% FBS for 2 hours cell lysate with HA722839 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA722839 at 1/2,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: HeLa serum starved for 40 hours then add 20% FBS for 2 hours cell lysate (input)
Lane 2: HA722839 IP in HeLa serum starved for 40 hours then add 20% FBS for 2 hours cell lysate
Lane 3: Rabbit IgG instead of HA722839 in HeLa serum starved for 40 hours then add 20% FBS for 2 hours cell lysate

Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 59 seconds; ECL: K1801