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CD14

CD14 is a glycoprotein primarily expressed on the surface of macrophages and monocytes, and to a lesser extent by dendritic cells and neutrophils. As a multifunctional lipopolysaccharide-binding protein, CD14 plays a key role as a co-receptor for several Toll-like receptors (TLRs) facilitating in the detection of bacterial lipopolysaccharides (LPS). In the presence of lipopolysaccharide-binding proteins (LBP), CD14 molecules can bind LPS which promotes LPS endocytosis and initiates the TRIF-dependent signaling pathway leading to the activation of vital transcription factors, such as the nuclear factor of activated T-cells (NFAT) transcription factors.

 

Table 1. CD14 At A Glance

Specification
Value
SynonymsLPS receptor, Monocyte differentiation antigen CD14, Myeloid cell-specific leucine-rich glycoprotein
Cellular localizationCell membrane
Cellular ExpressionLangerhans cells, Granulocytes, Monocytes, Macrophages
Receptor TypeGlycosylphosphatidylinositol (GPI)-anchored endotoxin receptor
LigandAffinity for bacterial lipopolysaccharide (LPS)
Functionco-receptor for Toll-like receptor (TLR) family
Associated Signaling PathwaysTLRs signaling pathway (upon LPS stimulation)


Types of CD14: mCD14 and sCD14


CD14 is present in two forms, either anchored to the cell surface or as a soluble CD14 protein found in circulation. Cell surface CD14, or mCD14, is anchored to the membrane by a glycosylphosphatidylinositol tail and is mainly distributed on the surface of mature macrophages, monocytes and neutrophils. This 55 kDa glycoprotein consists of a polypeptide chain approximately 356 amino acid residues long, which contains N-terminal residues at segments 39 to 44 that are essential for LPS binding. This interaction between LPS/LBP and CD14 results in the production of cytokines and the initiation of an immune response.

Soluble CD14, or sCD14, is somewhat smaller than that of mCD14, weighing ?48 kDa. It is produced by monocytes and appears in circulation either after shedding of mCD14 under the catalysis of proteases or it can be directly secreted by intracellular vesicles. Soluble CD14 competes with mCD14 for LPS binding and depending upon the concentration, normal or elevated, sCD14 can behave as either an LPS agonist or antagonist, respectively. In clinical diagnostics, elevated plasma levels of sCD14 are closely associated with HIV-related mortality.




CD14 Antibodies


Monoclonal antibodies, designed to recognize and react with the human CD14, are frequently used in flow cytometry and immunofluorescence applications to identify immune cell subsets expressing CD14. Cells known to express CD14 surface markers include macrophages, monocytes, neutrophils, dendritic cells, Langerhans cells and granulocytes

AAT Bioquest offers a comprehensive catalog of CD14 antibodies purified by affinity chromatography and conjugated to a variety of fluorophores under optimal conditions that minimize unconjugated fluorophore and antibody. Available fluorophores include:

  • iFluor™ dyes - bright, photostable dyes with optimized flow cytometry (FACS), fluorescence imaging and in vivo imaging applications.
  • mFluor™ dyes - bright, photostable dyes with optimized for flow cytometry (FACS) applications.
  • Alexa Fluor® dyes - suitable for flow cytometry (FACS) and fluorescence imaging applications.
  • Classic dyes - suitable for flow cytometry (FACS) and fluorescence imaging applications.
  • Phycobiliproteins and Tandem dyes - intensely bright dyes for flow cytometry (FACS) and multiparametric analysis.


Table 2. Overview of monoclonal anti-human CD14 antibodies.

Immunogen
Class
Species Reactivity
Host, Isotype
Clone
Applications¹
Recognizes human CD14MonoclonalHumanMouse, IgG1, kappa61D3FC, ELISA, IHC, IF, WB
Recognizes human CD14MonoclonalHumanMouse, IgMHI221FC, ELISA, IHC, IF, WB

Applications

  1. FC = Flow Cytometry; FN = Functional Assay; ELISA = Enzyme-linked immunosorbent assay; IF = Immunofluorescence; IHC = Immunohistochemistry; IHC-F = Immunohistochemistry (Frozen); ICC = Immunocytochemistry; WB = Western Blot; IP = Immunoprecipitation.



iFluor™ Dyes Labeled to CD14 Antibodies

The following table outlines the fluorescence properties of available iFluor™ dye labeled anti-human CD14 antibodies for use in flow cytometry (FACS), immunoblotting (WB) and fluorescence imaging applications. Conjugates made with iFluor™ dyes exhibit superior brightness and photostability, outperforming Alexa Fluor® conjugates and other spectrally similar conjugates. For additional information on iFluor™ dye-labeled CD14 antibodies and availability of other clones click on any label in the table below.



Table 3. Available iFluor™ dye-labeled CD14 antibody conjugates.

Label
Ex (nm)
Em (nm)
ε¹
Φ²
CF 260 nm³
CF 280 nm?
iFluor™ 35034444820,0000.950.830.23
iFluor™ 40540242537,0000.910.480.77
iFluor™ 43043349540,0000.780.680.3
iFluor™ 45045150240,0000.820.450.27
iFluor™ 48849151675,0000.90.210.11
iFluor™ 51452755480,0000.830.250.11
iFluor™ 53254356390,0000.680.260.16
iFluor™ 546541557100,0000.670.250.15
iFluor™ 555556569100,0000.640.230.14
iFluor™ 560559571130,0000.570.0480.069

Note

  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Φ = fluorescence quantum yield in aqueous buffer (pH 7.2).
  3. CF at 260 nm is the correction factor used for eliminating the dye contribution to the absorbance at 260 nm (for oligo and nucleic acid labeling).
  4. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).



mFluor™ Dyes Labeled to CD14 Antibodies

The following table outlines the fluorescence properties of available mFluor™ dye labeled anti-human CD14 antibodies for use in flow cytometry (FACS). Each mFluor™ dyes is optimally excited by one of the major laser lines commonly equipped in flow cytometers, such as the 405 nm, 488 nm, 532 nm, 561 nm or 633 nm laser lines. For additional information on mFluor™ dye-labeled CD14 antibodies and availability of other clones click on any label in the table below.



Table 4. Available mFluor™ dye-labeled CD14 antibody conjugates.

Label
Ex (nm)
Em (nm)
ε¹
Φ²
CF 260 nm³
CF 280 nm?
mFluor™ UV37535438835,0000.940.0990.138
mFluor™ Violet 45040644525,0000.920.3380.078
mFluor™ UV46036245615,0000.860.350.134
mFluor™ Violet 50042649735,0000.810.7690.365
mFluor™ Violet 51040950430,0000.860.4640.366
mFluor™ Violet 54040053215,0000.641.3920.529
mFluor™ Blue 570552564120,0000.080.2280.179
mFluor™ Green 62052161750,0000.060.8950.569
mFluor™ Yellow 630546625110,0000.010.2830.413
mFluor™ Red 700657694250,0000.0290.1350.127

Note

  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Φ = fluorescence quantum yield in aqueous buffer (pH 7.2).
  3. CF at 260 nm is the correction factor used for eliminating the dye contribution to the absorbance at 260 nm (for oligo and nucleic acid labeling).
  4. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).



Alexa Fluor® Dyes Labeled to CD14 Antibodies

The following table outlines the fluorescence properties of available Alexa Fluor® dye labeled anti-human CD14 antibodies for use in flow cytometry (FACS) and fluorescence imaging applications. For additional information on Alexa Fluor® dye-labeled CD14 antibodies and availability of other clones click on any label in the table below.



Table 5. Available Alexa Fluor® dye-labeled CD14 antibody conjugates.

Label
Ex (nm)
Em (nm)
ε¹
Φ²
CF 280 nm³
Alexa Fluor® 35034344119,000N/D0.19
Alexa Fluor® 48849952073,0000.920.11
Alexa Fluor® 555553568155,0000.10.08
Alexa Fluor® 59459061892,0000.660.56
Alexa Fluor® 647650671270,0000.330.03
Alexa Fluor® 680681704183,0000.360.05
Alexa Fluor® 700696719205,0000.250.07
Alexa Fluor® 750752776290,0000.120.04

Note

  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Φ = fluorescence quantum yield in aqueous buffer (pH 7.2).
  3. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).



Classic Dyes Labeled to CD14 Antibodies

The following table outlines the fluorescence properties of available classic dye labeled anti-human CD14 antibodies for use in flow cytometry (FACS) and fluorescence imaging applications. For additional information on classic dye-labeled CD14 antibodies and availability of other clones click on any label in the table below.



Table 6. Available FITC, TRITC and other classic dye-labeled CD14 antibody conjugates.

Label
Ex (nm)
Em (nm)
ε¹
Φ²
CF 280 nm³
Pacific Blue™40445546,0000.780.2
FITC49151673,0000.790.254
Pacific Orange™40055125,000N/D0.6
TRITC54457085,0000.10.34

Note

  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).
  2. Φ = fluorescence quantum yield in aqueous buffer (pH 7.2).
  3. CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptides and protein labeling).



PE, APC, PerCP and Tandem Dyes Labeled to CD14 Antibodies

The following table outlines the fluorescence properties of available phcoerythrin (PE), allophycocyanin (APC), PerCP and tandem dye labeled anti-human CD14 antibodies for use in flow cytometry (FACS). Phycobiliproteins are uncharacteristically bright due to their high molar extinction coefficients and quantum yields, an enviable quality when imaging low-abundance targets. However, since phycobiliprotiens photobleach rapidly, they are not recommended for microscopy. For additional information on phycobiliprotein-labeled CD14 antibodies and availability of other clones click on any label in the table below.



Table 7. Available PE, APC, PerCP and tandem dye-labeled CD14 antibody conjugates.

Label
Ex (nm)
Em (nm)
ε¹
Laser Line (nm)
PE5665741,960,000488 nm, 532 nm, 561-568 nm
PE-Texas Red®5676151,960,000488 nm, 532 nm, 561-568 nm
PE-Alexa Fluor® 6105676271,960,000488 nm, 532 nm, 561-568 nm
PE-Cy5®5656661,960,000488 nm, 532 nm, 561-568 nm
PE-iFluor™ 6475696661,960,000488 nm, 532 nm, 561-568 nm
PE-Alexa Fluor® 7005667211,960,000488 nm, 532 nm, 561-568 nm
PE-Cy7®5667781,960,000488 nm, 532 nm, 561-568 nm
PE-iFluor™ 7505667781,960,000488 nm, 532 nm, 561-568 nm
APC651660700,000633-647 nm
APC-iFluor™ 700651710700,000633-647 nm

Note

  1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm-1M-1).



References


  1. Clement, M., Ladell, K., Ekeruche-Makinde, J., Miles, J. J., Edwards, E. S., Dolton, G., Williams, T., Schauenburg, A. J., Cole, D. K., Lauder, S. N., Gallimore, A. M., Godkin, A. J., Burrows, S. R., Price, D. A., Sewell, A. K., & Wooldridge, L. (2011). Anti-CD14 antibodies can trigger CD14+ T cell effector function in the absence of TCR engagement and improve peptide-MHCI tetramer staining. Journal of immunology (Baltimore, Md. : 1950), 187(2), 654–663. https://doi.org/10.4049/jimmunol.1003941
  2. Hailman, E., Lichenstein, H. S., Wurfel, M. M., Miller, D. S., Johnson, D. A., Kelley, M., Busse, L. A., Zukowski, M. M., & Wright, S. D. (1994). Lipopolysaccharide (LPS)-binding protein accelerates the binding of LPS to CD14. The Journal of experimental medicine, 179(1), 269–277. https://doi.org/10.1084/jem.179.1.269
  3. Ranoa, D. R., Kelley, S. L., & Tapping, R. I. (2013). Human lipopolysaccharide-binding protein (LBP) and CD14 independently deliver triacylated lipoproteins to Toll-like receptor 1 (TLR1) and TLR2 and enhance formation of the ternary signaling complex. The Journal of biological chemistry, 288(14), 9729–9741. https://doi.org/10.1074/jbc.M113.453266
  4. Shive, C. L., Jiang, W., Anthony, D. D., & Lederman, M. M. (2015). Soluble CD14 is a nonspecific marker of monocyte activation. AIDS (London, England), 29(10), 1263–1265. https://doi.org/10.1097/QAD.0000000000000735
  5. Zanoni, I., & Granucci, F. (2013). Role of CD14 in host protection against infections and in metabolism regulation. Frontiers in cellular and infection microbiology, 3, 32. https://doi.org/10.3389/fcimb.2013.00032