Recombinant Anti-SPARC antibody [EPR25122-122] (ab290636)

Product name

Anti-SPARC antibody [EPR25122-122]
See all SPARC primary antibodies
Description

Rabbit monoclonal [EPR25122-122] to SPARC
Host species

Rabbit
Specificity

This antibody does not react with Rat species for IHC application.
Tested applications

Suitable for: IHC-P, ICC/IF, WB, Flow Cyt (Intra), IPmore details
Species reactivity

Reacts with: Mouse, Rat
Does not react with: Human
Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Mouse brain, heart, placenta tissue lysate. Rat brain, heart, placenta tissue lysates. C2C12, Neuro-2a, bEnd.3 and C6 whole cell lysate, untreated mouse and rat brain tissue lysate, mouse and rat brain tissue lysate treated with Protein Deglycosylation MIX II. Flow Cyt (intra): rat and mouse primary neuron cells. IP: Neuro-2a whole cell lysate, rat brain tissue. IHC-P: mouse hepatocellular, kidney, cerebrum tissues. ICC/IF: Permeabilized bEnd.3 and C6 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.

Form

Liquid
Storage instructions

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

pH: 7.20
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

EPR25122-122
Isotype

IgG
Research areas

Alternative Versions
- Anti-SPARC antibody [EPR25122-122] - BSA and Azide free (ab290647)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Related Buffer
- VeriBlot for IP Detection Reagent (HRP) (ab131366)
Related Products
- Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081)
- Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab290636 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application	Abreviews	Notes
IHC-P		1/5000.
ICC/IF		1/100.
WB		1/1000. Predicted molecular weight: 35 kDa.
Flow Cyt (Intra)		1/600.
IP		1/30.

Notes
IHC-P 1/5000.
ICC/IF 1/100.
WB 1/1000. Predicted molecular weight: 35 kDa.
Flow Cyt (Intra) 1/600.
IP 1/30.

Function

Appears to regulate cell growth through interactions with the extracellular matrix and cytokines. Binds calcium and copper, several types of collagen, albumin, thrombospondin, PDGF and cell membranes. There are two calcium binding sites; an acidic domain that binds 5 to 8 Ca(2+) with a low affinity and an EF-hand loop that binds a Ca(2+) ion with a high affinity.
Sequence similarities

Belongs to the SPARC family.
Contains 1 EF-hand domain.
Contains 1 follistatin-like domain.
Contains 1 Kazal-like domain.
Developmental stage

Expressed at high levels in tissues undergoing morphogenesis, remodeling and wound repair.
Cellular localization

Secreted > extracellular space > extracellular matrix > basement membrane. In or around the basement membrane.
Target information above from: UniProt accession P09486 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 20692 Mouse
- Entrez Gene: 24791 Rat
- SwissProt: P07214 Mouse
- SwissProt: P16975 Rat
- Unigene: 291442 Mouse
- Unigene: 98989 Rat
Alternative names
- AA517111 antibody
- Basement membrane protein 40 antibody
- Basement-membrane protein 40 antibody
- BM 40 antibody
- BM-40 antibody
- BM40 antibody
- Cysteine rich protein antibody
- hm:zeh0062 antibody
- MGC128090 antibody
- OI17 antibody
- ON antibody
- Osteonectin antibody
- Secreted acidic cystein rich glycoprotein antibody
- Secreted protein acidic and cysteine rich antibody
- Secreted protein acidic and rich in cysteine antibody
- Secreted protein acidic cysteine rich (osteonectin) antibody
- Secreted protein acidic cysteine rich antibody
- SPARC antibody
- SPRC antibody
- SPRC_HUMAN antibody
see all

Immunocytochemistry/ Immunofluorescence - Anti-SPARC antibody [EPR25122-122] (ab290636)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized bEnd.3 (mouse brain endothelioma cell line) cells labelling SPARC with primary antibody anti-SPARC (ab290636) at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150081) secondary antibody at 1/1000 dilution (2.0 µg/mL). Confocal image showing cytoplasmic staining in bEnd.3 cells. Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used to counterstain tubulin at 1/200 dilution (2.5 µg/mL). The nuclear counter stain is DAPI (blue).
The secondary antibody only control is : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (2.0 µg/mL).
Immunocytochemistry/ Immunofluorescence - Anti-SPARC antibody [EPR25122-122] (ab290636)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized C6 (rat glial tumour cell line) cells labelling SPARC with primary antibody anti-SPARC (ab290636) at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150081) secondary antibody at 1/1000 dilution (2.0 µg/mL). Confocal image showing cytoplasmic staining in C6 cells. Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used to counterstain tubulin at 1/200 dilution (2.5 µg/mL). The nuclear counter stain is DAPI (blue).
The secondary antibody only control is : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (2.0 µg/mL).
Western blot - Anti-SPARC antibody [EPR25122-122] (ab290636)

All lanes : Anti-SPARC antibody [EPR25122-122] (ab290636) at 1/1000 dilution

Lane 1 : Untreated mouse brain tissue lysate
Lane 2 : Mouse brain tissue lysate treated with Protein Deglycosylation MIX II
Lane 3 : Untreated rat brain tissue lysate
Lane 4 : Rat brain tissue lysate treated with Protein Deglycosylation MIX II

Lysates/proteins at 15 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 35 kDa
Observed band size: 40, 37 kDa why is the actual band size different from the predicted?

Exposure time: 103 seconds

Blocking and diluting buffer and concentration was 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with that described in the literature (PMID: 29449802; PMID: 9008236).
SPARC is a glycosylated protein and can be deglycosylated by Protein Deglycosylation MIX II.
The additional band is expected to be a SPARC cleavage product (PMID: 9008236).
Western blot - Anti-SPARC antibody [EPR25122-122] (ab290636)

All lanes : Anti-SPARC antibody [EPR25122-122] (ab290636) at 1/1000 dilution

Lane 1 : C2C12 (mouse myoblast) whole cell lysate
Lane 2 : bEnd.3 (mouse brain endothelioma) whole cell lysate
Lane 3 : Neuro-2a (mouse neuroblastoma) whole cell lysate
Lane 4 : C6 (rat glioma) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 35 kDa
Observed band size: 40, 37 kDa why is the actual band size different from the predicted?

Blocking and diluting buffer and concentration was 5% NFDM/TBST.

Exposure time: 3.25 seconds.

The expression profile/molecular weight observed is consistent with that described in the literature (PMID: 29449802; PMID: 9008236).
The additional band is expected to be a SPARC cleavage product (PMID: 9008236).
Western blot - Anti-SPARC antibody [EPR25122-122] (ab290636)

All lanes : Anti-SPARC antibody [EPR25122-122] (ab290636) at 1/1000 dilution

Lane 1 : Mouse brain tissue lysate
Lane 2 : Mouse heart tissue lysate
Lane 3 : Mouse placenta tissue lysate
Lane 4 : Rat brain tissue lysate
Lane 5 : Rat heart tissue lysate
Lane 6 : Rat placenta tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 35 kDa
Observed band size: 40, 37 kDa why is the actual band size different from the predicted?

Blocking and diluting buffer and concentration was 5% NFDM/TBST.

The expression profile/molecular weight observed is consistent with that described in the literature (PMID: 29449802; PMID: 9008236).
The additional band is expected to be a SPARC cleavage product (PMID: 9008236).

Exposure time:

Lane 2: 5.5 seconds
Lane 1, 3-6: 3.25 seconds
Immunoprecipitation - Anti-SPARC antibody [EPR25122-122] (ab290636)

SPARC was immunoprecipitated from rat brain tissue lysate with ab290636 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab290636 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: Rat brain tissue lysate 10 µg

Lane 2: ab290636 IP in Rat brain tissue lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab290636 in rat brain tissue lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 10 seconds
Immunoprecipitation - Anti-SPARC antibody [EPR25122-122] (ab290636)

SPARC was immunoprecipitated from Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate with ab290636 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab290636 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate 10 µg

Lane 2: ab290636 IP in Neuro-2a whole cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab290636 in Neuro-2a whole cell lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 10 seconds
Flow Cytometry (Intracellular) - Anti-SPARC antibody [EPR25122-122] (ab290636)

Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Rat primary neuron cells labelling SPARC with ab290636 at 1/600 dilution (0.1µg) (Right) compared with a Rabbit monoclonal IgG (ab172730) (Left). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.
Flow Cytometry (Intracellular) - Anti-SPARC antibody [EPR25122-122] (ab290636)

Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Mouse primary neuron cell cells labelling SPARC with ab290636 at 1/600 dilution (0.1µg) (Right) compared with a Rabbit monoclonal IgG (ab172730) (Left) isotype control 0. A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SPARC antibody [EPR25122-122] (ab290636)

Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labelling SPARC with ab290636 at 1/15000 (0.041 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) . Positive staining on microglia of mouse cerebrum. The section was incubated with ab290636 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) .

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-SPARC antibody [EPR25122-122] (ab290636)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labelling SPARC with ab290636 at 1/15000 (0.041 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) . Positive staining on glomerulus and endothelium of mouse kidney. The section was incubated with ab290636 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) .

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-SPARC antibody [EPR25122-122] (ab290636)

Immunohistochemical analysis of paraffin-embedded mouse hepatocellular tissue labelling SPARC with ab290636 at 1/15000 (0.041 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) . Positive staining on interstitial cells of mouse hepatocellular carcinoma. The section was incubated with ab290636 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) .

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
Anti-SPARC antibody [EPR25122-122] (ab290636)

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

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Key features and details

Overview

Product name

Description

Host species

Specificity

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Isotype control

Related Buffer

Related Products

Applications

The Abpromise guarantee

Target

Function

Sequence similarities

Developmental stage

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

References (0)

Customer reviews and Q&As