Anti-RecA antibody [EPR29094-561]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RecA antibody [EPR29094-561] (AB323376)

Immunohistochemical analysis of paraffin-embedded (A) HEK-293T (human embryonic kidney epithelial cell) transfected with a E.coli RecA expression vector containing a His-tag; (B) HEK-293T transfected with a Bacillus subtilis recA expression vector containing a His-tag; (C) HEK-293T transfected with a Staphylococcus aureus recA expression vector containing a His-tag; (D) HEK-293T transfected with a Helicobacter pylori recA expression vector containing a His-tag; (E) HEK-293T transfected with empty vector containing a His-tag. tissue labeling RecA with ab323376 at 1/1000 (0.503 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) HEK-293T transfected with a E.coli RecA expression vector containing a His-tag, no staining on (B) HEK-293T transfected with a Bacillus subtilis recA expression vector containing a His-tag, (C) HEK-293T transfected with a Staphylococcus aureus recA expression vector containing a His-tag, (D) HEK-293T transfected with a Helicobacter pylori recA expression vector containing a His tag, and (E) HEK-293T transfected with empty vector containing a His tag. The section was incubated with ab323376 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• I-ELISA

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Indirect ELISA - Anti-RecA antibody [EPR29094-561] (AB323376)

Indirect ELISA analysis of ab323376 at 1000-0 ng/ml. The Secondary antibody used was Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1 : 2500 dilution dilution.

Antigen : E.coli RecA recombinant protein.

Antigen concentration : 1000 ng/ml

• IP

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Immunoprecipitation - Anti-RecA antibody [EPR29094-561] (AB323376)

RecA was immunoprecipitated from 0.35 mg E.coli lysate with ab323376 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab323376 at 1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : E.coli lysate
Lane 2 : ab323376 IP in E.coli lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab323376 in E.coli lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST

All lanes:

Immunoprecipitation - Anti-RecA antibody [EPR29094-561] (ab323376) at 1/30 dilution

All lanes:

E.coli lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 3s

• WB

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Western blot - Anti-RecA antibody [EPR29094-561] (AB323376)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This antibody does weak cross-react with Bacillus subtilis RecA, but does not cross-react with Staphylococcus aureus RecA and Helicobacter pylori RecA .

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-RecA antibody [EPR29094-561] (ab323376) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a His-tag, whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a E.coli RecA expression vector containing a His-tag, whole cell lysate at 20 µg

Lane 3:

293T cells transfected with a Bacillus subtilis RecA expression vector containing a His-tag, whole cell lysate at 20 µg

Lane 4:

293T cells transfected with a Staphylococcus aureus RecA expression vector containing a His-tag, whole cell lysate at 20 µg

Lane 5:

293T cells transfected with a Helicobacter pylori RecA expression vector containing a His-tag, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 35-45 kDa,124 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-RecA antibody [EPR29094-561] (AB323376)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-RecA antibody [EPR29094-561] (ab323376) at 1/1000 dilution

Lane 1:

No-tag E.coli RecA recombinant protein at 10 ng

Lane 2:

E.coli lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 40 kDa

false

Exposure time: 10s