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CD68/SR-D1 Antibody (FA-11) - BSA Free

Catalog # NBP2-33337 | Novus Biologicals a Bio-Techne Brand
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NBP2-33337
NBP2-33337SS

Key Product Details

Species Reactivity

Mouse

Applications

Flow (Intracellular), Flow Cytometry, Functional, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry-Frozen, Immunohistochemistry-Paraffin, Immunoprecipitation, Single Cell Western, Western Blot

Label

Unconjugated

Antibody Source

Monoclonal Rat IgG2A Clone # FA-11

Format

BSA Free

Concentration

1.0 mg/ml

Product Summary for CD68/SR-D1 Antibody (FA-11) - BSA Free

Immunogen

This CD68/SR-D1 Antibody (FA-11) was developed against purified ConA acceptor glycoproteins from the P815 cell line.

Clonality

Monoclonal

Host

Rat

Isotype

IgG2A

Scientific Data Images for CD68/SR-D1 Antibody (FA-11) - BSA Free

Immunohistochemistry-Paraffin: CD68/SR-D1 Antibody (FA-11) [NBP2-33337] - Mouse spleen cryosection.
Immunohistochemistry-Paraffin: CD68/SR-D1 Antibody (FA-11) [NBP2-33337] - Mouse spleen.
Immunocytochemistry/Immunofluorescence: CD68/SR-D1 Antibody (FA-11) [NBP2-33337] - Wehi-3 cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X TBS + 0.5% Triton X-100. The cells were incubated with anti CD68 (FA-11) [NBP2-33337] at a 1:100 dilution overnight at 4C and detected with an anti-rat DyLight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:200 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Applications for CD68/SR-D1 Antibody (FA-11) - BSA Free

Application
Recommended Usage

Flow Cytometry

1:50-1:100

Functional

reported in scientific literature (PMID 11085350)

Immunocytochemistry/ Immunofluorescence

1:10-1:500. Use reported in scientific literature (PMID 34478932)

Immunohistochemistry

1:10-1:500. Use reported in scientific literature (PMID 34478932)

Immunohistochemistry-Frozen

1:10-1:500

Immunohistochemistry-Paraffin

1:10-1:500

Immunoprecipitation

1:10-1:500

Western Blot

1:100-1:2000
Application Notes
For Flow Cytometry: Use 10 ul of suggested dilution to label 10^6 cells in 100 ul. IHC requires antigen retrieval using heat treatment prior to staining of paraffin sections. Sodium citrate buffer pH 6.0 is recommended for this purpose.
Please Note: Optimal dilutions of this antibody should be experimentally determined.

Published Applications

Read 37 publications using NBP2-33337 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: CD68/SR-D1

Human CD68, also known as GP110, LAMP4, Scavenger Receptor D1 (SR-D1) or macrosialin in mouse, encodes a 110-kD transmembrane glycoprotein that belongs to the lysosomal/endosomal-associated membrane glycoprotein (LAMP) family. Members of the LAMP family include LAMP-1, LAMP-2, dendritic cell (DC)-LAMP (aka CD208), and brain and dendritic cell-associated (BAD)-LAMP (aka LAMP-5). Unlike the two LAMP domains facing the lysosomal lumen in LAMP-1 and LAMP-2, CD68 has a single LAMP domain containing four cystines spaced 36-37 residues apart along with an N-terminal Mucin-like domain. The 354 amino acid (a.a.) human CD68 and 326 a.a. murine ortholog share 80.6% a.a. sequence identity (1).

CD68 is highly expressed in cells of the mononuclear phagocyte system such as macrophages, microglia, osteoclasts, and myeloid dendritic cells (DCs); and is expressed to a lesser extent in lymphoid cells (CD19+ B lymphocytes and CD4+ T lymphocytes), human umbilical cord mesenchymal stem cells (MSCs), fibroblasts, endothelial cells, multiple non-hematopoietic cancer cell lines, and human arterial intimal smooth muscle cells (SMCs). Expression has been also observed in diseased states for granulocytes and neutrophils, in particular basophils from myeloproliferative disorders and intestinal neutrophils from inflammatory bowel disease (IBD), respectively (1).

Although the function of CD68 has yet to be established, it has often been used as an immunohistochemistry (IHC) marker of inflammation and for granular cell tumors (GCTs). CD68+ tumor associated macrophages (TAMs) has been suggested to be a predictive marker for poor cancer prognosis, but a meta-analysis showed the presence of CD68 is not correlated with survival (2). In addition, a role in hepatic malaria infection has been reported based on the finding that peptide P39 binds CD68, considered a receptor for malaria sporozoite, and inhibits parasite entry into Kupffer cells. CD68 was deemed a member of the Scavenger receptor family due to its upregulation in macrophages following inflammatory stimuli, ability to bind modified LDL, phosphatidylserine, and apoptotic cells, as well as shuttling between the plasma membrane and endosomes. CD68 has been linked to atherogenesis based on binding and internalization of its ligand, oxLDL (1).

References

1. Chistiakov, DA, Killingsworth, MC, Myasoedova, VA. Orekhov AN, Bobryshev YV. (2017) CD68/macrosialin: not just a histochemical marker. Lab Invest. 97:4-13. PMID: 27869795

2. Troiano G, Caponio VCA, Adipietro I, Tepedino M, Santoro R, Laino L, Lo Russo L, Cirillo N, Lo Muzio L. (2019) Prognostic significance of CD68+ and CD163+ tumor associated macrophages in head and neck squamous cell carcinoma: A systematic review and meta-analysis. Oral Oncol. 93:66-75. PMID: 31109698.

Alternate Names

CD68, gp110, Macrosialin, SCARD1, SR-D1, SRD1

Entrez Gene IDs

12514 (Mouse)

Gene Symbol

CD68

UniProt

Product Documents for CD68/SR-D1 Antibody (FA-11) - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot number in the search box below.

Product Specific Notices for CD68/SR-D1 Antibody (FA-11) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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Protocols

View specific protocols for CD68/SR-D1 Antibody (FA-11) - BSA Free (NBP2-33337):

Protocol for Flow Cytometry Intracellular Staining
Sample Preparation.
1. Grow cells to 60-85% confluency. Flow cytometry requires between 2 x 105 and 1 x 106 cells for optimal performance.
2. If cells are adherent, harvest gently by washing once with staining buffer and then scraping. Avoid using trypsin as this can disrupt certain epitopes of interest. If enzymatic harvest is required, use Accutase, Collagenase, or TrypLE Express for a less damaging option.
3. Reserve 100 uL for counting, then transfer cell volume into a 50 mL conical tube and centrifuge for 8 minutes at 400 RCF.
a. Count cells using a hemocytometer and a 1:1 trypan blue exclusion stain to determine cell viability before starting the flow protocol. If cells appear blue, do not proceed.
4. Re-suspend cells to a concentration of 1 x 106 cells/mL in staining buffer (NBP2-26247).
5. Aliquot out 100 uL samples in accordance with your experimental samples.

Tip: When cell surface and intracellular staining are required in the same sample, it is advisable that the cell surface staining be performed first since the fixation and permeabilization steps might reduce the availability of surface antigens.

Intracellular Staining.
Tip: When performing intracellular staining, it is important to use appropriate fixation and permeabilization reagents based upon the target and its subcellular location. Generally, our Intracellular Flow Assay Kit (NBP2-29450) is a good place to start as it contains an optimized combination of reagents for intracellular staining as well as an inhibitor of intracellular protein transport (necessary if staining secreted proteins). Certain targets may require more gentle or transient permeabilization protocols such as the commonly employed methanol or saponin-based methods.
Protocol for Cytoplasmic Targets:
1. Fix the cells by adding 100 uL fixation solution (such as 4% PFA) to each sample for 10-15 minutes.
2. Permeabilize cells by adding 100 uL of a permeabilization buffer to every 1 x 106 cells present in the sample. Mix well and incubate at room temperature for 15 minutes.
a. For cytoplasmic targets, use a gentle permeabilization solution such as 1X PBS + 0.5% Saponin or 1X PBS + 0.5% Tween-20.
b. To maintain the permeabilized state throughout your experiment, use staining buffer + 0.1% of the permeabilization reagent (i.e. 0.1% Tween-20 or 0.1% Saponin).
3. Following the 15 minute incubation, add 2 mL of the staining buffer + 0.1% permeabilizer to each sample.
4. Centrifuge for 1 minute at 400 RCF.
5. Discard supernatant and re-suspend in 100 uL of staining buffer + 0.1% permeabilizer.
6. Add appropriate amount of each antibody (eg. 1 test or 1 ug per sample, as experimentally determined).
7. Mix well and incubate at room temperature for 30 minutes- 1 hour. Gently mix samples every 10-15 minutes.
8. Following the primary/conjugate incubation, add 1-2 mL/sample of staining buffer +0.1% permeabilizer and centrifuge for 1 minute at 400 RCF.
9. Wash twice by re-suspending cells in staining buffer (2 mL for tubes or 200 uL for wells) and centrifuging at 400 RCF for 5 minutes. Discard supernatant.
10. Add appropriate amount of secondary antibody (as experimentally determined) to each sample.
11. Incubate at room temperature in dark for 20 minutes.
12. Add 1-2 mL of staining buffer and centrifuge at 400 RCF for 1 minute and discard supernatant.
13. Wash twice by re-suspending cells in staining buffer (2 mL for tubes or 200 uL for wells) and centrifuging at 400 RCF for 5 minutes. Discard supernatant.
14. Resuspend in an appropriate volume of staining buffer (usually 500 uL per sample) and proceed with analysis on your flow cytometer.

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