MA5-12596

antibody from Invitrogen Antibodies

Targeting: KRT5 EBS2, KRT5A

Provider product page for MA5-12596

Western blot

Antibody data

Antibody Data
Antigen structure
References [42]
Comments [0]
Validations
Western blot [3]
Immunocytochemistry [2]
Immunohistochemistry [3]
Flow cytometry [1]
Other assay [10]

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Product number: MA5-12596 - Provider product page
Provider: Invitrogen Antibodies
Product name: Cytokeratin 5 Monoclonal Antibody (XM26)
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Description: MA5-12596 targets Cytokeratin 5 in IHC (P) applications and shows reactivity with Human samples.
Antibody clone number: XM26
Concentration: Conc. Not Determined

KRT5 protein structure - MA5-12596 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on Whole cell extract (30 µg lysate) of A431. The blot was probed with Anti-Cytokeratin 5 Mouse Monoclonal Antibody (Product # MA5-12596, 1:250 dilution) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). A ~60 kDa band corresponding to Cytokeratin 5 was observed in the cell line tested. Known quantity of protein sample was electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of Cytokeratin 5 was achieved by transfecting A-431 with Cytokeratin 5 specific siRNAs (Silencer® select Product # s7956, S7957). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Cytokeratin 5 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Cytokeratin 5 Monoclonal Antibody (XM26) (Product # MA5-12596, 1:1000) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to Cytokeratin 5.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-Cytokeratin 5 Monoclonal Antibody (XM26) (Product # MA5-12596) and a 60kDa band corresponding to Cytokeratin 5 was observed across the cell lines tested except MCF7, IMR-32 and SH-SY5Y along with non specific bands around 45-55 kDa. Whole cell extracts (30 µg lysate) of MCF 10A (Lane 1), MCF7 (Lane 2), A-431 (Lane 3), HaCaT (Lane 4), IMR-32 (Lane 5), SH-SY5Y (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Cytokeratin 5 was performed using 70% confluent log phase A-431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Cytokeratin 5 (XM26) Mouse Monoclonal Antibody (Product # MA5-12596) at 1:250 dilution in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Cytokeratin 5 was performed using 70% confluent log phase A-431 cells. The cells were fixed with ice-cold acetone at 4°C for 5 minutes and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with Cytokeratin 5 Monoclonal Antibody (XM26) (Product # MA5-12596) at 1:200 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175), (1:2000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300 dilution). Panel d represents the merged image showing cytoskeletal localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Formalin-fixed, paraffin-embedded human mesothelioma stained with Keratin 5 antibody using peroxidase-conjugate and AEC chromogen. Note cytoplasmic staining of tumor cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Cytokeratin 5 was performed on cryosections of human skin tissue. Tissues were blocked in 10% normal goat serum in 1X PBS containing 0.1% Triton X-100 (1X PBS-T) for 1 hour at room temperature (RT). The tissues were labeled with a Cytokeratin 5 monoclonal antibody (clone XM26, green, Product # MA5-12596) diluted 1:50 in 3% normal goat serum in 1X PBS-T for 1 hour at RT, followed by detection with a Goat anti-Mouse IgG1, Alexa Fluor 488 secondary antibody (Product # A-21121) diluted 1:2000 in 3% normal goat serum in 1X PBS-T for 1 hour at RT. Nuclei (blue) were stained with DAPI, included in ProLong Gold Anti-Fade Mountant (Product # P36931). Images were taken on an inverted microscope at 20X magnification. Data courtesy of Dr. Jiyoon Lee at Indiana University School of Medicine.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of KRT5 was performed using formalin-fixed paraffin-embedded human skin (malignant melanoma) tissue sections. To expose the target protein, heat-induced epitope retrieval was performed on de-paraffinized sections using eBioscience™ IHC Antigen Retrieval Solution - Low pH (10X) (Product # 00-4955-58) diluted to 1X solution in water in a decloaking chamber at 110 degree Celsius for 15 minutes. Following antigen retrieval, the sections were blocked with 2% normal goat serum in 1X PBS for 45 minutes at room temperature and then probed with or without Cytokeratin 5 Monoclonal Antibody (XM26) (Product # MA5-12596) at 1:100 dilution in 0.1% normal goat serum overnight at 4 degree Celsius in a humidified chamber. Detection was performed using Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32723) at a dilution of 1:2000 in 0.1% normal goat serum for 45 minutes at room temperature. ReadyProbes™ Tissue Autofluorescence Quenching Kit (Product # R37630) was used to quench autofluorescence from the tissues. Nuclei were stained with DAPI (Product # D1306) and the sections were mounted using ProLong™ Glass Antifade Mountant (Product # P36984). The images were captured on EVOS™ M7000 Imaging System (Product # AMF7000) at 20X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIG 7 SARS-CoV-2 infects ciliated and goblet epithelial cells but not basal and club cells. Epithelial cells of the mock- and SARS-CoV-2-infected HAE-ALI B9-20 cultures at 4 dpi (MOI = 0.2) were dissociated from the Transwell insert and cytospun onto slides. The cells on the slides were fixed, permeabilized, and immunostained with anti-NP and together with anti-beta-tubulin IV (A), and anti-MUC5AC (B), anti-cytokeratin 5 (C), and anti-SCGB1A1 (D), respectively. Confocal images were taken at a magnification of x63. Nuclei were stained with DAPI (blue).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 Delivery of CCL2-neutralizing antibodies did not significantly affect macrophage infiltration or tumor angiogenesis. (A) Primary tumor tissues were co-immunofluorescent stained for CK5 (green) and F4/80 (red). Secondary antibody controls overlaid with 4',6-diamidino-2-phenylindole are shown. (B) Primary tumor tissues were immunostained for vWF8. Magnified insert shows positive staining. Staining was quantified by Image J. Statistical analysis was performed using two-tailed Student's t test. Statistical significance was determined by P < .05. ns = not significant. Mean +- SEM is shown. Scale bar = 200 mum. Figure 5

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 3 Figure Treatment of TRb-expressing MDA-MB-468 cells with T 3 represses cytokeratin genes. (A) Gene Set Enrichment Analysis performed between T 3 -treated 468-EV and 468-TRbeta expression analysis revealed Gene Ontology Biological Processes or Hallmark gene sets involving differentially expressed cytokeratins genes (adjusted p value < 1e-7). (B) Heatmap of all expressed KRT genes in 468-EV or 468-TRbeta cells following Vehicle or 10 nM T 3 treatment. (C) Normalized gene expression bar plots of the cytokeratin genes KRT5 and KRT14 from RNA-seq data, n = 3 for each condition. (D) and (E) Protein expression of KRT5 and KRT14 were evaluated by western blot, compared with beta-Actin as a loading control. The blot presented is representative of three independent experiments. **** p < 0.0001. TRbeta, thyroid hormone receptor beta [Color figure can be viewed at wileyonlinelibrary.com ]

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Keratin 5 and fibulin-4 were found in the amyloid deposits of Bowen's carcinoma and lichen amyloidosis skin. (a) The skin from a Bowen's carcinoma (patient 1) was stained with anti-fibulin-4, anti-keratin 5, and anti-fibrillin-1 antibodies (the left column) together with thioflavin T. Bar = 50 mum. ( b ) Paraffin-embedded sections (patient 2, lichen amyloidosis) were stained with anti-fibulin-4 (red) and anti-keratin 5 (green, data not shown) and with anti-keratin 5 and thioflavin T. Bar = 100 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fibulin-4 and keratin 5 were found in the amyloid deposits of lichen amyloidosis (patient 3). ( a ) Consecutive frozen sections were stained with anti-keratin 5 or with anti-fibulin-4, and amyloid deposits were detected with thioflavin T. Each merged image is also shown. ( b ) The staining with anti-fibulin-4 in the same section as the second row of a was shown at lower magnification. This image also shows the localization of fibulin-4 in normal human skin. Fibulin-4 localizes in elastic fibers and in the surrounding blood vessels, not only in the papil1ary dermis but also in the reticular dermis. Bar = 100 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 Prolactin suppression of BCL6 and subsequently CK5 is mediated by the Stat5 pathway (A) Immunoblots of BCL6, CK5, phosphorylated-Stat5 (p-Stat5), Stat5a, and GAPDH protein levels in extracts of T47D cultures exposed to adenovirus carrying either Stat5a or LacZ control for 24 h and subsequently treated with vehicle (Cntrl), Prolactin (PRL), beta-Estradiol (E2), or R5020 (Pg) for 48 h. (B) Immunoblots of BCL6, CK5, phosphorylated-Stat5 (p-Stat5), Stat5, Jak2, and GAPDH protein levels in extracts of T47D cultures treated with non-target shRNA (NTC) or Jak2 shRNA for 48h, followed by addition of vehicle (Cntrl), Prolactin (PRL), beta-Estradiol (E2), or R5020 (Pg) for 48 h. (C) Immunoblots of BCL6, phosphorylated-ERK1/2 (p-ERK1/2), ERK1/2, and GAPDH protein levels in extracts of T47D cultures pre-treated with Pg for 24 h followed by incubation with or without prolactin (PRL) in the presence or absence of Mek inhibitor (U0126) for 3 and 6 h. (D) Immunoblots of BCL6, phosphorylated-Akt (p-Akt), Akt, and GAPDH protein levels in extracts of T47D cultures pre-treated with Pg for 24 h followed by incubation with or without prolactin (PRL) in the presence or absence of PI3K inhibitor (Ly294002) for 3 and 6 h.