Anti-KIFC1 antibody [11445]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KIFC1 antibody [11445] (AB172620)

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue laneling KIFC1 with ab172620 (unpurified) at 1/50 dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-KIFC1 antibody [11445] (AB172620)

Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling KIFC1 with purified ab172620 at 1/50 dilution (4.0 μg/ml). Cells were fixed in 100% Methanol. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 (2.5 μg/ml) dilution. 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

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-KIFC1 antibody [11445] (AB172620)

Immunofluorescent staining of HeLa cells labeling KIFC1 with ab172620 (unpurified) at 1/50 dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KIFC1 antibody [11445] (AB172620)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human stomach tissue sections labeling KIFC1 with purified ab172620 at 1/16000 dilution (0.01 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-KIFC1 antibody [11445] (AB172620)

Intracellular Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling KIFC1 with purified ab172620 at 1/20 dilution (10μg/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).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KIFC1 antibody [11445] (AB172620)

Immunohistochemical analysis of paraffin-embedded Human cervical carcinoma tissue labeling KIFC1 with ab172620 (unpurified) at 1/50 dilution.

• IP

Unknown

Immunoprecipitation - Anti-KIFC1 antibody [11445] (AB172620)

ab172620 (purified) at 1/20 dilution (1ug) immunoprecipitating KIFC1 in Jurkat whole cell lysates.
Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysates 10ug
Lane 2 (+) : ab172620 & Jurkat whole cell lysates
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab172620 in Jurkat whole cell lysates
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-KIFC1 antibody [11445] (ab172620)

Predicted band size: 74 kDa

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• WB

Supplier Data

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

Western blot analysis on immunoprecipitation pellet from 293T cell lysate, labeling KIFC1 with ab172620 (unpurified) at 1/10 dilution.

All lanes:

Western blot - Anti-KIFC1 antibody [11445] (ab172620)

Predicted band size: 74 kDa

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• WB

Supplier Data

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : KIFC1 knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HeLa whole cell lysate (20 μg)
Lane 4 : HepG2 whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab172620 (purified) observed at 74 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab172620 was shown to specifically react with KIFC1 in wild-type cells as signal was lost in KIFC1 knockout cells. Wild-type and KIFC1 knockout samples were subjected to SDS-PAGE. ab172620 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 0.228 μg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-KIFC1 antibody [11445] (ab172620)

Predicted band size: 74 kDa

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• WB

Lab

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

Western blot : Rabbit Monoclonal[11445] to KIFC1 ab172620 staining at 1/50000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 75 kDa in Wild-type A549 cell lysates with no signal observed at this size in KIFC1 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

Lanes 1 - 3:

Western blot - Anti-KIFC1 antibody [11445] (ab172620) at 1/50000 dilution

Lanes 1 - 4:

Western blot - Anti-PERK antibody [EPR19876-294] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/perk-antibody-epr19876-294-bsa-and-azide-free-ab254249'>ab254249</a>) at 1/50000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

KIFC1 knockout A549 at 20 µg

Lane 2:

KIFC1 knockout A549 at 20 g

Lane 3:

U-87 MG at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 74 kDa

Observed band size: 75 kDa

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• WB

Supplier Data

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

All lanes:

Western blot - Anti-KIFC1 antibody [11445] (ab172620) at 1/10000 dilution

Lane 1:

HeLa lysate at 10 µg

Lane 2:

293T lysate at 10 µg

Lane 3:

HepG2 lysate at 10 µg

Predicted band size: 74 kDa

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• WB

Lab

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

All lanes:

Western blot - Anti-KIFC1 antibody [11445] (ab172620) at 1/50000 dilution

All lanes:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 15 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: 74 kDa

Observed band size: 74 kDa

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• WB

CiteAb

Western blot - Anti-KIFC1 antibody [11445] (AB172620)

KIFC1 western blot using anti-KIFC1 antibody [11445] ab172620. Publication image and figure legend from She, Z. Y., Pan, M. Y., et al., 2017, Oncotarget, PubMed 28430595.

ab172620 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 ab172620 please see the product overview.

Depletion of KIFC1 led to the disorganization and dispersal of the Golgi apparatus(A, B) HEK293T cells were transfected with the shLuc, shkifc1-1, shkifc1-2, shkifc1-3 shRNA plasmids for 90 hr and analyzed by immunofluorescence assays, respectively. shLuciferase was used for the control group. DAPI (blue), GFP (green), GM130 (red), DIC (Differential Interference Contrast). Scale bars, 5 μm. (C) Representative three dimensional images of the Golgi apparatus (GM130, red) in the shRNA transfected HEK293T cells. The zoom is the magnified view of the indicated box. Scale bars, 5 μm. (D, E) Western Blot analysis of RNAi knockdown efficiency in HEK293T cells after 90 hr. Five different kifc1 targeting shRNA plasmids were used as candidates. The first, second and fourth plasmid (1, 2, 4 designated as shkifc1-1, shkifc1-2 and shkifc1-3, respectively) were selected for this study. GAPDH was used as the loading control. (F) Quantification the ratios of cells containing normal Golgi apparatus in the transfected HEK293T cells after kifc1 knockdown assay for 90 hr. Data are represented as mean ± SEM. ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test). (G) Quantification of the ratios of disorganized Golgi apparatus (gray color) and the dispersed Golgi (black color) into vesicles after s after kifc1 knockdown assay for 90 hr. Data are represented as mean ± SEM. ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test).

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