Anti-hnRNP K antibody [F45 P9 C7]

• IHC-P

AbReview20480****

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

ab23644 staining hnRNP K in Human head and neck tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 1 hour at 25°C; antigen retrieval was by heat mediation with Tris EDTA (pH9). Samples were incubated with primary antibody (1 µg/ml in TBST) for 16 hours at 4°C. An undiluted HRP-conjugated Rabbit anti-mouse polyclonal was used as the secondary antibody.

This image is courtesy of an Abreview submitted by Satyendra Tripathi

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

IHC image of hnRNP K staining in a section of formalin-fixed paraffin-embedded normal human colon* performed on a Leica BOND™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab23644, 1ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

Overlay histogram showing HeLa cells stained with ab23644 (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 followed by the antibody (ab23644, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 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 with 0.1% PBS-Tween for 20 min used under the same conditions.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

Formalin-fixed, paraffin embedded tissue sections from human normal colon or human colon carcinoma stained with ab23644 mouse monoclonal to hnRNP K. hnRNP K expression is increased in the tumour tissue.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

ab23644 staining hnRNP K in HeLa cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab23644 at 1µg/ml and ab6046, Rabbit polyclonal to beta Tubulin - Loading Control. Cells were then incubated with ab150117, Goat polyclonal Secondary Antibody to Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150080, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 594) at 1/1000 dilution (shown in pseudocolour magenta). Nuclear DNA was labelled with DAPI (shown in blue). Also suitable in cells fixed with 100% methanol (5 min).Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• WB

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Western blot - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

All lanes:

Western blot - Anti-hnRNP K antibody [F45 P9 C7] (ab23644) at 5 µg/mL

Lane 1:

NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 20 µg

Lane 2:

MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 20 µg

Lane 3:

Testis (Mouse) Tissue Lysate at 20 µg

Lane 4:

Lung (Mouse) Tissue Lysate at 20 µg

Lane 5:

PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG H&L (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-hrp-preadsorbed-ab97040'>ab97040</a>) at 1/5000 dilution

Predicted band size: 50 kDa

Observed band size: 21 kDa,24 kDa,54 kDa,56 kDa

true

Exposure time: 4min

• WB

Ap237728****

Western blot - Anti-hnRNP K antibody [F45 P9 C7] (AB23644)

Lane 1:

Marker

Lanes 2 - 5:

Western blot - Anti-hnRNP K antibody [F45 P9 C7] (ab23644) at 1 µg/mL

Lane 2:

HeLa (Human epithelial carcinoma cell line) Nuclear Lysate (ab27251) at 20 µg

Lane 3:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab27252) at 20 µg

Lane 4:

Western blot - Jurkat whole cell lysate (<a href='/en-us/products/cell-lysates/jurkat-whole-cell-lysate-ab7899'>ab7899</a>) at 20 µg

Lane 5:

Western blot - HEK-293 whole cell lysate (<a href='/en-us/products/cell-lysates/hek-293-whole-cell-lysate-ab7902'>ab7902</a>) at 20 µg

Secondary

All lanes:

Western blot - Rabbit Anti-Mouse IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/rabbit-mouse-igg-h-l-hrp-ab6728'>ab6728</a>) at 1/10000 dilution

Predicted band size: 50 kDa

false