Name: Anti-Collagen I antibody [EPR24331-53]
SKU: ab270993
Rating: 1 (1 reviews)

Anti-Collagen I antibody [EPR24331-53] is a rabbit recombinant monoclonal antibody that is used to detect Collagen I in Flow cytometry (Intra), ICC/IF, IHC-Fr, IHC-P, IP, Western blot, mIHC. Suitable for Mouse, Rat samples.

- Using biophysical QC, antibody identity is confirmed at a molecular level for unrivaled batch-batch consistency
- Validated on the Leica BOND™ RX automated IHC staining platform for Collagen I IHC
- Cited in over 70 publications

Related conjugates and formulations

ab279711
Carrier free
Anti-Collagen I antibody [EPR24331-53] - BSA and Azide free
ab313153
Conjugated
Alexa Fluor® 555 Anti-Collagen I antibody [EPR24331-53]
ab310848
Conjugated
APC Anti-Collagen I antibody [EPR24331-53]
ab310918
Conjugated
PE Anti-Collagen I antibody [EPR24331-53]
ab313351
Conjugated
Alexa Fluor® 594 Anti-Collagen I antibody [EPR24331-53]
ab312943
Conjugated
Alexa Fluor® 568 Anti-Collagen I antibody [EPR24331-53]
ab309248
Conjugated
Alexa Fluor® 488 Anti-Collagen I antibody [EPR24331-53]
ab309367
Conjugated
Alexa Fluor® 647 Anti-Collagen I antibody [EPR24331-53]
ab321535
Conjugated
Alexa Fluor® 750 Anti-Collagen I antibody [EPR24331-53]

Images

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (AB270993), expandable thumbnail

Publications

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

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Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	ICC/IF	IP	WB	IHC-Fr	Flow Cyt (Intra)	IHC-P	mIHC
Human	Not recommended	Not recommended	Not recommended	Not recommended	Not recommended	Not recommended	Not recommended
Mouse	Tested	Tested	Tested	Tested	Tested	Tested	Tested
Rat	Expected	Expected	Tested	Tested	Expected	Tested	Expected

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/2000	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/30	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/1000	Notes For mouse samples, please use high sensitivity substrate.
Species Rat	Dilution info 1/1000	Notes For mouse samples, please use high sensitivity substrate.

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes For mouse samples, please use high sensitivity substrate.

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/100	Notes Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20)
Species Rat	Dilution info 1/100	Notes Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20)

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/500	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/500	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Rat	Dilution info 1/500	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/500	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Target data

See full target information Col1a1

Function

Type I collagen is a member of group I collagen (fibrillar forming collagen).

Alternative names

Cola1, Col1a1, Collagen alpha-1(I) chain, Alpha-1 type I collagen

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Clone number: EPR24331-53
Purification technique: Affinity purification Protein A
Specificity: FURTHER INFORMATION ON SPECIFICITY (Chinese Version) available under the support & downloads section.
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage duration: 1-2 weeks
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: -20°C
Aliquoting information: Upon delivery aliquot
Storage information: Avoid freeze / thaw cycle

Notes

Anti-Collagen I antibody [EPR24331-53] (ab270993) was developed by Abcam using patented rabbit monoclonal antibody technology and is validated for use in Flow Cyt (Intra), ICC/IF, IHC-Fr, IHC-P, IP and WB.

Abcam's high quality manufacturing and validation processes ensure Anti-Collagen I antibody [EPR24331-53] (ab270993) has high sensitivity and specificity alongside high lot-to-lot consistency and reproducibility.

Anti-Collagen I antibody [EPR24331-53] (ab270993) specifically detects Collagen I (UniProt ID: P11087; Molecular weight: 95kDa) and is sold in a convenient trial size to enable initial testing (20 µL) and larger sizes for subsequent scaling up experiments (100 µL and 1 mL).

Conjugation-ready, carrier free format available for antibody clone EPR24331-53 - Anti-Collagen I antibody [EPR24331-53] - BSA and Azide free ab279711.

Antibody clone EPR24331-53 is also available pre-conjugated to a variety of labels for your convenience - Alexa Fluor^® 488, Alexa Fluor^® 647, APC, PE, Alexa Fluor^® 568, Alexa Fluor^® 555, Alexa Fluor^® 594, Alexa Fluor^® 750 (Alexa Fluor® 488 Anti-Collagen I antibody [EPR24331-53] ab309248, Alexa Fluor® 647 Anti-Collagen I antibody [EPR24331-53] ab309367, APC Anti-Collagen I antibody [EPR24331-53] ab310848, PE Anti-Collagen I antibody [EPR24331-53] ab310918, Alexa Fluor® 568 Anti-Collagen I antibody [EPR24331-53] ab312943, Alexa Fluor® 555 Anti-Collagen I antibody [EPR24331-53] ab313153, Alexa Fluor® 594 Anti-Collagen I antibody [EPR24331-53] ab313351, Alexa Fluor® 750 Anti-Collagen I antibody [EPR24331-53] ab321535).

Collagen I is a major component of the tumor microenvironment, influencing cancer progression by enhancing tumor stiffness, promoting metastasis and inhibiting anti-tumor immune responses. It also affects the efficacy of immunotherapies, making it a critical factor in cancer prognosis and treatment strategies.

Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

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 batch production

For more information, read more on recombinant antibodies.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.

Activity summary

Collagen type I also called collagen I is a structural protein expressed mainly in connective tissues such as skin tendon bone and ligaments. It serves as an important component in providing mechanical strength and integrity to these tissues. Collagen I is a fibrillar collagen known for its triple-helix structure composed of two alpha-1 chains and one alpha-2 chain and has a molecular mass of approximately 300 kDa. Researchers often employ collagen western blot and collagen ELISA techniques for its detection. Collagen suppliers offer various collagen antibodies used in these assays to study its distribution and function.

Biological function summary

Collagen type I plays a central role in maintaining the extracellular matrix and supporting cellular environments. It interacts with other matrix proteins and cells forming complexes that help in tissue development and repair. Type I collagen is especially important in bone matrix working alongside minerals like hydroxyapatite to provide rigidity and support. Anti-collagen antibodies aid in studying its biological functions and interactions which are critical to understanding tissue dynamics.

Pathways

Collagen type I interacts with multiple signaling cascades involved in tissue remodeling and repair. It is a significant player in the TGF-β pathway which regulates fibrosis and wound healing processes. In these pathways proteins such as fibronectin and integrins work in concert with collagen type I to orchestrate cellular responses to damage. Researchers often examine its role in these pathways to uncover therapeutic possibilities for disease interventions.

Associated diseases and disorders

Collagen type I has strong connections to conditions like osteogenesis imperfecta and fibrosis. Mutations or irregularities in collagen I production can lead to osteogenesis imperfecta a genetic disorder characterized by brittle bones. In fibrosis excessive collagen deposition disrupts normal tissue architecture contributing to organ dysfunction. In both conditions type I collagen interacts with other proteins like matrix metalloproteinases which modulate its breakdown and remodeling highlighting its importance in disease pathology.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

16 product images

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of paraffin-embedded Mouse pancreatic cancer tissue labeling Collagen I with ab270993 at 1/500 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining in connective tissues of mouse pancreatic cancer. The section was incubated with ab270993 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Blocking buffer and concentration: 5% NFDM/TBST
Diluting buffer and concentration: 5% NFDM/TBST
We are unsure how to define these extra bands below 100kDa.
The molecular weight observed is consistent with what has been described in the literature (PMID:23940311;PMID:29853175).
All lanes: Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993) at 1/1000 dilution
All lanes: NIH/3T3 (mouse embryonic fibroblast), whole cell lysate at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 139 kDa
Observed band size: 220 kDa
Exposure time: 20s
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of paraffin-embedded Mouse skin tissue labeling Collagen I with ab270993 at 1/500 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining in connective tissues of mouse skin. The section was incubated with ab270993 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunoprecipitation - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Collagen I was immunoprecipitated from 0.35 mg Mouse skin tissue lysate 10 ug with ab270993 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab270993 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
Lane 1: Mouse skin tissue lysate 10 ug
Lane 2: ab270993 IP in Mouse skin tissue lysate
Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab270993 in Mouse skin tissue lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 32 seconds
All lanes: Immunoprecipitation - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Predicted band size: 139 kDa
Observed band size: 138 kDa
Immunocytochemistry/ Immunofluorescence - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized NIH/3T3 cells labelling Collagen I with ab270993 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in NIH/3T3 cells is observed. Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
Immunohistochemistry (Frozen sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse skin tissue labeling Collagen I with ab270993 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Positive staining on mouse skin is observed. The nuclear counterstain was DAPI (Blue).
Secondary antibody control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488)at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
Flow Cytometry (Intracellular) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (Mouse embryonic fibroblast) cells labelling Collagen I with ab270993 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at 1/2000 dilution was used as the secondary antibody.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of paraffin-embedded Mouse stomach tissue labeling Collagen I with ab270993 at 1/500 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining in connective tissues of mouse stomach. The section was incubated with ab270993 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Blocking and diluting buffer and concentration: 5% NFDM/TBST
The observed MW is consistent with what has been described in the literature (PMID: 27740527;PMID: 22278938; PMID: 26973392).
Exposure time: Lane 1: 3.25 seconds
Lane 2: 5.5 seconds
All lanes: Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993) at 1/1000 dilution
Lane 1: Mouse skin tissue lysate at 20 µg
Lane 2: Rat skin tissue lysate at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 139 kDa
Observed band size: 138 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of paraffin-embedded Rat skin tissue labeling Collagen I with ab270993 at 1/500 dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining in connective tissues of rat skin. The section was incubated with ab270993 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunohistochemistry (Frozen sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat skin tissue labeling Collagen I with ab270993 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Positive staining on rat skin is observed. The nuclear counterstain was DAPI (Blue).
Secondary antibody control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488)at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Blocking buffer and concentration : 5% NFDM/TBST.

Diluting buffer and concentration : 5% NFDM/TBST.

Anti-GAPDH antibody [EPR16891] - Loading Control ab181602 was used as a loading control.

This blot was produced using a high sensitivity ECL substrate.
All lanes: Western blot - Anti-Collagen I antibody [EPR24331-53] (ab270993) at 1/1000 dilution
Lane 1: Mouse skin tissue lysate at 20 µg
Lane 2: Mouse kidney tissue lysate at 20 µg
Lane 3: Mouse lung tissue lysate at 20 µg
Lane 4: Mouse heart tissue lysate at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 139 kDa
Observed band size: 139 kDa
Exposure time: 100s
Multiplex immunohistochemistry - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Mouse skin staining of Cytokeratin 1 + Cytokeratin 2e with Anti-Cytokeratin 1 + Cytokeratin 2e antibody [EPR27257-91] ab315228 at a 1/2000 (0.263 μg/ml) dilution, Collagen I with ab270993 and Langerin with Anti-Langerin antibody [EPR24685-12] ab283686 at a 1/1000 dilution followed by a ready to use secondary antibody Opal Polymer HRP Ms + Rb.
Panel A: merged staining of anti-Cytokeratin 1 + Cytokeratin 2e (green; Opal™520), anti-Collagen I (magenta; Opal™690) and anti-Langerin (gray; Opal™570) on mouse skin.

Panel B: anti-Cytokeratin 1 + Cytokeratin 2e staining epidermis keratin layer in mouse skin.

Panel C: anti-Collagen I staining connective tissues in mouse skin.

Panel D: anti-Langerin staining Langerhans cells in mouse skin.

Nuclear DNA was labeled with DAPI (shown in blue).
The section was incubated in three rounds of staining: in the order of Anti-Cytokeratin 1 + Cytokeratin 2e antibody [EPR27257-91] ab315228, ab270993 and Anti-Langerin antibody [EPR24685-12] ab283686 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system,
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of formalin fixed paraffin embedded mouse skin labelling Collagen I with ab270993 at a concentration of 0.025µg/ml. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with a Bond™ Polymer Refine Detection kit. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution 2) for 20mins.
ab270993 anti-Collagen I antibody [EPR24331-53] was incubated for 15mins at room temperature. Sections were counterstained with Hematoxylin. Image inset shows absence of staining in secondary antibody only control.
Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Immunohistochemical analysis of formalin fixed paraffin embedded mouse skin labelling Collagen I with ab270993 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a ChromoMap DAB kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32mins.
ab270993 anti-Collagen I antibody [EPR24331-53] was incubated for 16mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.
Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)
Multiplex immunohistochemistry - Anti-Collagen I antibody [EPR24331-53] (ab270993)
Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse thyroid staining of Thyroid Peroxidase/TPO with Anti-Thyroid Peroxidase/TPO antibody [EPR23574-405] ab278525 at a 1/5000 (0.109 μg/ml) dilution, PAX8 with Anti-PAX8 antibody [EPR23508-20] ab239363 at 1/2000 (0.254 μg/ml) dilution, and Collagen I with ab270993 at a 1/500 (1.152 μg/ml) dilution, followed by a ready to use secondary antibody Opal Polymer HRP Ms + Rb.
Panel A: merged staining of anti-Thyroid peroxidase (green; Opal™ 520), anti-PAX8 (magenta; Opal™ 690) and anti-Collagen I (gray; Opal™ 570) on mouse thyroid.

Panel B: anti-Thyroid peroxidase showed cytoplasmic and membranous staining on mouse thyroid.

Panel C: anti-PAX8 showed nucleus staining on mouse thyroid.

Panel D: anti-Collagen I staining connective tissues in mouse thyroid.

Nuclear DNA was labeled with DAPI (shown in blue).
The section was incubated in three rounds of staining: in the order of Anti-Thyroid Peroxidase/TPO antibody [EPR23574-405] ab278525, Anti-PAX8 antibody [EPR23508-20] ab239363 and ab270993 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.