IL-6 antibody

Catalog No. ABIN2689779

Product Details anti-IL-6 Antibody

Target: IL-6 (IL6) (Interleukin 6 (IL6))

Reactivity: Human

Host: Rat

Clonality: Monoclonal

Conjugate: This IL-6 antibody is un-conjugated

Application: Blocking Antibody (Inhibition)

Brand: BD Pharmingen™

Characteristics: The MQ2-6A3 antibody reacts with human interleukin-6 (IL-6). The immunogen used to generate the MQ2-6A3 hybridoma was recombinant human IL-6. This is a neutralizing antibody. PBMC were isolated from human peripheral blood by density gradient centrifugation and were cultured with PMA (Sigma, Cat. No. P-8139, 5 ng/mL) and ionomycin (Sigma, Cat. #I-0634, 500 ng/mL) in the presence of GolgiStop™ (Cat. No. 554724) overnight at 37 °C. The activated cells were harvested and the presence of IL-6 producing cells was detected by immunocytochemistry using a three -step staining procedure that employs a Biotin Goat anti-Rat IgG secondary antibody (Cat. No. 559286) and a horseradish peroxidase-based detection system. To demonstrate specificity of staining the binding of the MQ2-6A3 (Cat. No. 559068) antibody was blocked by the preincubation of the purified antibody with excess recombinant human IL-6 protein (Cat. No. 550071, data not shown). (Nomarski optics, original magnification 400X).

BD Pharmingen™ Purified Rat Anti-Human IL-6 - Purified - Clone MQ2-6A3 - Isotype Rat IgG2a, κ - Reactivity Hu - 0.25 mg

Purification: The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Immunogen: Recombinant human IL-6

Clone: MQ2

Isotype: IgG2a kappa

Application Details

Application Notes: Optimal working dilution should be determined by the investigator.

Assay Procedure:

FOR IMMUNOCYTOCHEMICAL STAINING OF SINGLE-CELL PREPARATIONS This procedure describes the immunoenzymatic technique of staining cytokines within individual cells that are immobilized on microscopic slides via adherence (adherent slides) or centrifugation (cytospins). ADHESION SLIDES 1. Harvest cells and wash them twice in PBS using centrifugation (400 x g for 5 min) to remove residual protein. 2. Adjust the cell concentration at 4 x 10^6 to 5 x 10^6 cells/mL in PBS. 3. Place 20 μL of the cell suspension in each well of the adhesion slides and let them adhere at room temperature (RT) for 20 min. Please note that the slides should be washed in PBS at RT for 5 min before transferring the cells. 4. Fix cells on slides using fixation buffer for 15 min at RT. 5. Wash slides 2X in PBS with 5 min incubations. 6. Block slides with PBS supplemented with 1 % (w/v) BSA (Sigma, Cat. No. A43-78) for 30 min at RT or 10 min at 37 °C. 7. Wash slides 2X in PBS and proceed with staining or air dry them and store them at -80 °C for future use. 8. Incubate slides with 20 μL of 1 % goat serum and PBS with 0.1 % (w/v) saponin for 30 min at RT. 9. Wash slides 2X with PBS with 5 min incubations. 10. Block endogenous peroxidase activity with Endogenous Peroxidase Blocking Buffer (20 μL/well) for 10 min at RT. 11. Wash 2X in PBS with 5 min incubations. 12. Incubate each well with Avidin (20 μL/well) for 15 min. 13. Wash 2X in PBS with 5 min incubations. 14. Incubate each well with Biotin (20 μL/well) for 15 min. 15. Wash 2X in PBS with 5 min incubations. 16. Incubate each well for 1 hr at RT with 20 μL of purified cytokine-specific antibody or appropriate immunoglobulin isotype control diluted in Pharmingen's IHC Diluent (Cat. No. 559148) supplemented with saponin. 17. Wash slides 2X in PBS with 5 min incubations. 18. Incubate each well with 20 μL of a biotinylated secondary antibody diluted in IHC Diluent for 30 min at RT. 19. Wash 2X in PBS with 5 min incubations. 20. Apply 20 μL of Streptavidin . HRP (Cat. No. 550946) to each well on slides and incubate for 30 min at RT. 21. Wash slides 2X with PBS with 5 minutes incubations. 22. Incubate with 3-3'-Diaminobenzidine tetra hydrochloride (DAB), (Cat. No. 550880) for less than 5 min at RT. 23. Stop the development of the color reaction by washing with PBS. 24. The slides are subsequently mounted in short-term storage mounding medium. CYTOSPINS 1. Assemble the Cytospin's sample chamber (e.g. Cytospin 3, Shandon, UK or comparable centrifuge), filter card, slide and cytospin racks according to manufacturer's specifications. 2. Load 40 μL of approximately 1 x 10e6 cells to each sample chamber. 3. Spin slides at 600 rpm for 2 min. 4. Take slides out of the cytospin rack and place them on a staining rack. 5. For fixation and staining please follow the steps 4 through 24 specified above for staining cells on adhesion slides.

Restrictions: For Research Use only

Handling

Concentration: 0.5 mg/mL

Buffer: Aqueous buffered solution containing ≤0.09 % sodium azide.

Preservative: Sodium azide

Precaution of Use: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Storage: 4 °C

Storage Comment: Store undiluted at 4°C.

References for anti-IL-6 antibody (ABIN2689779)

Abrams: "Immunoenzymetric assay of mouse and human cytokines using NIP-labeled anti-cytokine antibodies." in: Current protocols in immunology / edited by John E. Coligan ... [et al.], Vol. Chapter 6, pp. Unit 6.20, (2008) (PubMed).

Gaines Das, Poole: "The international standard for interleukin-6. Evaluation in an international collaborative study." in: Journal of immunological methods, Vol. 160, Issue 2, pp. 147-53, (1993) (PubMed).

Abrams, Roncarolo, Yssel, Andersson, Gleich, Silver: "Strategies of anti-cytokine monoclonal antibody development: immunoassay of IL-10 and IL-5 in clinical samples." in: Immunological reviews, Vol. 127, Issue 9-10, pp. 5-24, (1992) (PubMed).

Hsu, Raine, Fanger: "Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures." in: The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society, Vol. 29, Issue 4, pp. 577-80, (1981) (PubMed).

Target Details for IL-6

Target: IL-6 (IL6) (Interleukin 6 (IL6))

Alternative Name: IL-6 (IL6 Products)

Synonyms: BSF2 antibody, HGF antibody, HSF antibody, IFNB2 antibody, IL-6 antibody, Il-6 antibody, ILg6 antibody, Ifnb2 antibody, il6 antibody, CHIL-6 antibody, interleukin 6 antibody, interleukin-6 antibody, IL6 antibody, Il6 antibody, il-6 antibody, IL-6 antibody

Pathways: TLR Signaling, Hormone Transport, Negative Regulation of Hormone Secretion, Myometrial Relaxation and Contraction, Positive Regulation of Immune Effector Process, Production of Molecular Mediator of Immune Response, Regulation of Carbohydrate Metabolic Process, Autophagy, Cell RedoxHomeostasis, Cancer Immune Checkpoints, Inflammasome