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82944
CD14 (61D3) Mouse mAb (violetFluor™ 450 Conjugate)
Antibody Conjugates

CD14 (61D3) Mouse mAb (violetFluor™ 450 Conjugate) #82944

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Flow cytometric analysis of live human peripheral blood mononuclear cells using CD14 (61D3) Mouse mAb (violetFluor™ 450 Conjugate) (solid line) compared to concentration-matched Mouse (MOPC-21) mAb IgG1 Isotype Control (violetFluor™ 450 Conjugate) #40282 (dashed line). Analysis was performed on cells in the monocyte gate.

To Purchase # 82944S
Product # Size Price
82944S
500 µl  (100 tests) $ 285

Supporting Data

REACTIVITY H
SENSITIVITY Endogenous
MW (kDa)
Isotype Mouse IgG1

Product Description

This Cell Signaling Technology antibody is conjugated to violetFluor™ 450 and tested in-house for direct flow cytometry analysis in human cells.

Product Usage Information

Application Dilutions
Flow Cytometry 1:20

Storage:

Supplied in 10 mM NaH2PO4, 150 mM NaCl, 0.09% NaN3, 0.1% gelatin, pH7.2. This product is stable for 6 months when stored at 4ºC. Do not aliquot the antibody. Protect from light. Do not freeze.

Protocol

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Flow Cytometry

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.

  1. 20X Phosphate Buffered Saline (PBS): (#9808) To prepare 1 L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix.
  2. Incubation Buffer: Dissolve 0.5 g Bovine Serum Albumin (BSA) (#9998) in 100 ml 1X PBS. Store at 4°C.

B. Immunostaining

  1. Count cells using a hemacytometer or alternative method.
  2. Aliquot 0.5-1x106 cells into each assay tube (by volume).
  3. Add 2-3 ml Incubation Buffer to each tube and rinse by centrifugation. Repeat.
  4. Resuspend cells in 100 µl of diluted primary antibody (prepared in incubation buffer at the recommended dilution).
  5. Incubate for 1 hr at room temperature.
  6. Wash by centrifugation in 2-3 ml incubation buffer.
  7. Resuspend cells in the appropriate volume of PBS and analyze on flow cytometer.

posted June 2017

Protocol Id: 1504

Specificity / Sensitivity

CD14 (61D3) Mouse mAb (violetFluor™ 450 Conjugate) recognizes endogenous levels of total CD14 protein. This antibody detects an epitope within the extracellular domain.

Species Reactivity:

Human

Source / Purification

This monoclonal antibody was purified from tissue culture supernatant via affinity chromatography. The purified antibody was conjugated under optimal conditions, with unreacted dye removed from the preparation.

Background

CD14 is a leucine-rich repeat-containing pattern recognition receptor with expression largely restricted to the monocyte/macrophage cell lineage (1). Research studies have shown that CD14 is a bacterial lipopolysaccharide (LPS) binding glycoprotein, expressed as either a GPI-linked membrane protein or a soluble plasma protein (2). LPS induces an upregulation of GPI-linked CD14 expression, which facilitates TLR4 signaling and macrophage activation in response to bacterial infection (3-5).

The 61D3 antibody is widely used to identify cells of the monocyte/macrophage lineage (6).

  1. Wright, S.D. et al. (1991) J Exp Med 173, 1281-6.
  2. Ziegler-Heitbrock, H.W. and Ulevitch, R.J. (1993) Immunol Today 14, 121-5.
  3. Suzuki, M. et al. (2009) J Immunol 182, 6485-93.
  4. Pugin, J. et al. (1994) Immunity 1, 509-16.
  5. Zanoni, I. et al. (2011) Cell 147, 868-80.
  6. Flora, P.K. and Gregory, C.D. (1994) Eur J Immunol 24, 2625-32.

Pathways & Proteins

Explore pathways + proteins related to this product.

For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
violetFluor is a registered trademark of Tonbo Biosciences.