Antibodies, Proteins, Kits and Reagents for Life Science

345 091 ₸ Product size

100 µl 345 091 ₸

Order now and get it on Tuesday March 09, 2021

Anti-Collagen VI antibody [EPR17072] (ab182744)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR17072] to Collagen VI
Suitable for: WB, ICC/IF, IHC-P, mIHC
Knockout validated
Reacts with: Mouse, Rat, Human

Product name

Anti-Collagen VI antibody [EPR17072]
See all Collagen VI primary antibodies
Description

Rabbit monoclonal [EPR17072] to Collagen VI
Host species

Rabbit

Tested Applications & Species

Application	Species
ICC/IF	Human
IHC-P	Mouse Rat Human
mIHC	Human
WB	Mouse Human

See all applications and species data

Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Human skeletal muscle, Human placenta, Human fetal brain, Human fetal heart, Human fetal kidney, Human fetal spleen, Mouse heart, Mouse kidney, Mouse spleen, Rat kidney and Rat spleen lysates; HEK293T, WI-38 and NIH/3T3 whole cell lysates. IHC-P: Human liver, Human cardiac muscle, Mouse kidney and Rat stomach tissues. mIHC: Human liver. ICC/IF: HeLa cells.
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.
Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR17072
Isotype

IgG
Research areas

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen VI antibody [EPR17072] (ab182744)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling Collagen VI with ab182744 at 1/250 dilution, followed by prediluted HRP Polymer for Rabbit/Mouse IgG. Positive staining around sinusoidal endothelial basement membranes is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Collagen VI antibody [EPR17072] (ab182744)

All lanes : Anti-Collagen VI antibody [EPR17072] (ab182744) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : COL6A1 knockout HEK293T cell lysate
Lane 3 : Human skeletal muscle tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 109 kDa
Observed band size: 136 kDa
why is the actual band size different from the predicted?

Lanes 1-3: Merged signal (red and green). Green - ab182744 observed at 136 kDa. Red - loading control ab8245 observed at 36 kDa.

ab182744 Anti-Collagen VI antibody [EPR17072] was shown to specifically react with Collagen VI antibody in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab265060 (knockout cell lysate ab256879) was used. Wild-type and Collagen VI antibody knockout samples were subjected to SDS-PAGE. ab182744 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-Collagen VI antibody [EPR17072] (ab182744)

All lanes : Anti-Collagen VI antibody [EPR17072] (ab182744)

Lane 1 : Wild-type HEK293 whole cell lysate
Lane 2 : COL6A1 knockout HEK-293 whole cell lysate
Lane 3 : Human Skeletal Muscle whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 109 kDa

Lanes 1 - 3: Merged signal (red and green). Green - ab182744 observed at 109 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab182744 was shown to recognize Collagen VI in wild-type HEK293 cells as signal was lost at the expected MW in COL6A1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and COL6A1 knockout samples were subjected to SDS-PAGE. Ab182744 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution 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.
Multiplex immunohistochemistry - Anti-Collagen VI antibody [EPR17072] (ab182744)

Multiplex immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human liver tissue.

Panel A: Merged staining of Collagen VI (ab182744; green), anti-CD68 (ab213363; red) and anti-Lamin B1 (ab229025; magenta).

Panel B: Anti-Collagen VI (green) stained on extracellular matrix.

Panel C: Anti-CD68 (red) stained on Kupffer cells.

Panel D: Anti-Lamin B1 (magenta) stained on nuclear envelope.

Key protocol steps: The section was incubated in three rounds of staining with ab182744 (1/1000 dilution), ab213363 (1/1000 dilution) and ab229025 (1/4000 dilution) for 30 mins at room temperature. Each round was followed by tyramide signal amplification with the appropriate fluorophore.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

DAPI was used as a nuclear counter stain. A ready-to-use anti-Rabbit and Mouse Polymer HRP was used as a secondary.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-Collagen VI antibody [EPR17072] (ab182744)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling Collagen VI with ab182744 at 1/200 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/400 dilution (green). Cytoplasm staining on HeLa cell line is observed. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows:-
-ve control1 - ab182744 at 1/200 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2 - ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/400 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen VI antibody [EPR17072] (ab182744)

Immunohistochemical analysis of paraffin-embedded Human cardiac muscle tissue labeling Collagen VI with ab182744 at 1/250 dilution, followed by prediluted HRP Polymer for Rabbit/Mouse IgG. Positive staining on Human cardiac sarcolemma and interstitium is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen VI antibody [EPR17072] (ab182744)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labeling Collagen VI with ab182744 at 1/250 dilution, followed by prediluted HRP Polymer for Rabbit/Mouse IgG. Positive staining around basement membranes of Mouse renal tubules is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen VI antibody [EPR17072] (ab182744)

Immunohistochemical analysis of paraffin-embedded Rat stomach tissue labeling Collagen VI with ab182744 at 1/250 dilution, followed by prediluted HRP Polymer for Rabbit/Mouse IgG. Positive staining around Rat gastric epithelial basement membranes is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Collagen VI antibody [EPR17072] (ab182744)

All lanes : Anti-Collagen VI antibody [EPR17072] (ab182744) at 1/20000 dilution

Lane 1 : Human skeletal muscle lysate
Lane 2 : WI-38 (Human fetal lung fibroblast cells) whole cell lysate
Lane 3 : Human placenta lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 109 kDa
Observed band size: 147 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.
Due to posttranslational modifications, observed MW is greater than the predicted MW.
Western blot - Anti-Collagen VI antibody [EPR17072] (ab182744)

All lanes : Anti-Collagen VI antibody [EPR17072] (ab182744) at 1/2000 dilution

Lane 1 : Human fetal brain lysate
Lane 2 : Human fetal heart lysate
Lane 3 : Human fetal kidney lysate
Lane 4 : Human fetal spleen lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 109 kDa
Observed band size: 147 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.
Due to posttranslational modifications, observed MW is greater than the predicted MW.
Western blot - Anti-Collagen VI antibody [EPR17072] (ab182744)

All lanes : Anti-Collagen VI antibody [EPR17072] (ab182744) at 1/2000 dilution

Lane 1 : Mouse heart lysate
Lane 2 : Mouse kidney lysate
Lane 3 : Mouse spleen lysate
Lane 4 : Rat kidney lysate
Lane 5 : Rat spleen lysate
Lane 6 : NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 109 kDa
Observed band size: 147 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.
Due to posttranslational modifications, observed MW is greater than the predicted MW.
Anti-Collagen VI antibody [EPR17072] (ab182744)

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