Anti-PRC1 antibody [EP1513Y]

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-PRC1 antibody [EP1513Y] (AB51248)

Intracellular Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling PRC1 with Purified 51248 at 1/30 dilution (10 μg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-PRC1 antibody [EP1513Y] (AB51248)

Immunocytochemistry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling PRC1 with Purified 238427 at 1 : 50 dilution (5.06 μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1 : 200 (2.5 μg/ml). Goat anti rabbit IgG (Alexa Fluor® 488,ab150077) was used as the secondary antibody at 1 : 1000 (2 μg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• ICC/IF

AbReview13261****

Immunocytochemistry/ Immunofluorescence - Anti-PRC1 antibody [EP1513Y] (AB51248)

ab51248 staining PRC1 in human breast cancer cells by ICC/IF. The cells were paraformaldehyde fixed and blocked in 1% serum for 1 hour at 37°C without permeation step. The primary antibody was diluted 1/100 (PBS) and incubated with sample for 1 hour at 20°C. An Alexa Fluor® 488 conjugated donkey polyclonal to rabbit IgG, diluted 1/200 was used as secondary.

This image is courtesy of an anonymous Abreview.

• WB

Unknown

Western blot - Anti-PRC1 antibody [EP1513Y] (AB51248)

ab51248 recognise two isoforms of PRC1. But we are unsure how to define the extra bands at ~50kDa.

All lanes:

Western blot - Anti-PRC1 antibody [EP1513Y] (ab51248) at 1/1000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

C6 (Rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 3:

C2C12 (Mouse myoblasts myoblast) whole cell lysate at 20 µg

Lane 4:

Mouse kidney lysate at 20 µg

Lane 5:

Rat kidney 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 1/20000 dilution

Predicted band size: 72 kDa

false

• WB

CiteAb

Western blot - Anti-PRC1 antibody [EP1513Y] (AB51248)

PRC1 western blot using anti-PRC1 antibody [EP1513Y] ab51248. Publication image and figure legend from Dwivedi, D., Kumari, A., et al., 2019, J Cell Biol, PubMed 30674580.

ab51248 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab51248 please see the product overview.

Hook2 is required for p150glued and MKLP1 interaction during cytokinesis. (A) Lysates of HEK293T cells synchronized in either G2 phase or cytokinesis were IP with control IgG or anti-MKLP1 antibody and IB with indicated antibodies. (B) Ratio of normalized band intensity (cytokinesis lane) of IP p150glued and DIC to MKLP1 in A (n = 2). (C) Whole-cell lysates of HEK293T cells harvested during the cytokinesis stage were incubated with either MBP or MBP-tagged Hook2 N427 (WT, Q143A, and I150A) and IB for MKLP1. (D) Ratio of band intensity of pulldown to input Hook2 (WT/mutants) signal in C (n = 2). (E) Lysates of HEK293T cells treated with control or p150glued siRNA and harvested during cytokinesis stage were IP with control IgG or anti-Hook2 antibody and IB with the indicated antibodies. (F) Ratio of normalized band intensity (control siRNA) of coIP proteins (as indicated) to MKLP1 in E (n = 2). (G) Lysates of HEK293T cells transfected with indicated siRNA and harvested during cytokinesis stage were tested for dynactin association with MKLP1. (H) Ratio of normalized band intensity (control siRNA) of IP p150glued and DIC to MKLP1 in G (n = 3). (I) Representative images of HeLa cells ectopically expressing HA-tagged zebrafish Hook2 (HA-zHook2) costained for centrosomes, dynein, and dynactin. Bars, 10 μm; insets, 2 μm. (J) Lysates from asynchronous HEK293T cells treated with indicated siRNA and transfected with indicated plasmids were IP with control IgG or anti-DIC antibody and IB for p150glued. (K) Ratio of normalized band intensity (control siRNA) of IP p150glued to DIC in J (n = 2). (L) Representative differential interference contrast images of zebrafish embryos injected with either nontargeting control morpholino or indicated concentration of zHook2 morpholino immediately after fertilization and imaged at the indicated time. Each concentration of morpholino was injected in 100 fertilized embryos and monitored over time. Bars, 500 μm. Data represent mean ± SD (ns, not significant; ***, p < 0.001; Student's t test). (M) Proposed model depicting the role of Hook2 in mediating formation of dynein–dynactin complex and targeting of MKLP1 to the spindle midzone.

false