Human CD14 Quantikine QuicKit ELISA Summary
Sample Type & Volume Required Per Well
Cell Culture Supernates (50 uL), Serum (10 uL), EDTA Plasma (10 uL), Heparin Plasma (10 uL)
Assay Range
188.0 - 12,000 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma)
Specificity
Natural and recombinant human CD14.
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
Interference
No significant interference observed with available related molecules.
Sample Values
Serum/Plasma - Samples from apparently healthy volunteers were evaluated for the presence
of human CD14 in this assay. No medical histories were available for the donors used in this
study.
| Sample Type | Mean (ng/mL) | Range (ng/mL) |
| Serum (n=10) | 1901 | 1457-2260 |
| EDTA plasma (n=10) | 1534 | 1040-2015 |
| Heparin plasma (n=10) | 1678 | 1330-2480 |
Cell Culture Supernates - Human peripheral blood mononuclear cells (PBMCs)
(1 x 106
cells/mL) were cultured in RPMI supplemented with 10% fetal bovine serum,
2 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin sulfate. An aliquots of the
cell culture supernates was removed after 1 day, assayed for human CD14, and measured
6561 pg/mL.
Product Summary
The Quantikine® QuicKit™ Human CD14 Immunoassay is a one step, 80-minute solid phase ELISA designed to measure human CD14 levels in cell culture supernates, serum, and plasma. It contains CHO cell-expressed recombinant human CD14 and antibodies raised against the recombinant protein. Results obtained for naturally occurring human CD14 showed linear curves that were parallel to the standard curves obtained using the recombinant QuicKit™ standards. These results indicate that this kit can be used determine relative mass values for natural CD14.
Precision
Intra-Assay Precision (Precision within an assay) Two samples of known concentration were tested twenty times on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Two samples of known concentration were tested in ten separate assays to assess inter-assay precision. Assays were performed by at least three technicians
Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma
|
Intra-Assay Precision |
Inter-Assay Precision |
| Sample |
1 |
2 |
1 |
2 |
| n |
20 |
20 |
10 |
10 |
| Mean (pg/mL) |
1228 |
8177 |
1282 |
8360 |
| Standard Deviation |
28.3 |
262 |
77.4 |
339 |
| CV% |
2.3 |
3.2 |
6 |
4.1 |
Recovery
The recovery of human CD14 spiked to three different levels in samples throughout the range of the assay in various matrices was evaluated.
| Sample Type |
Average % Recovery |
Range % |
| Cell Culture Supernates (n=4) |
97 |
92-100 |
Linearity
To assess the linearity of the assay, samples containing high concentrations of human CD14 were diluted with calibrator diluent to produce samples with values within the dynamic range of the assay.
Scientific Data
HIV-1 Gag p24 Standard Curve
HIV-1 Gag p24 Standard Curve
Human CD14 Standard Curve
Product Datasheets
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Preparation and Storage
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: CD14
CD14 is a glycoprotein that mediates the interaction of
lipopolysaccharide (LPS, endotoxin)
with cells, thereby signaling the presence of gram-negative
bacteria (1-3). CD14 is either
soluble (CD14) (4, 5) or membrane-bound (mCD14) by a glycosylphosphatidylinositol
(GPI)
anchor (6, 7). mCD14 is a 55 kDa glycoprotein (1), while
CD14 varies from about 43 to 53 kDa,
depending on the degree of glycosylation and whether it was
synthesized without the anchor
or was shed by phospholipase cleavage of the anchor or by
proteolysis (12-14). There is no
evidence for different mRNAs for m- and CD14. There is no
apparent sequence homology with
other proteins. The sequence of human CD14 is 63-73%
identical to that of mouse, rat, or rabbit
CD14 (15).
mCD14 is expressed primarily on myeloid cells, such as
monocytes, macrophages and
neutrophils (1-3), the cells most sensitive to LPS, and to a
lesser extent on other cells, such as B
cells (8) and a circulating dendritic cell progenitor (9).
CD14 appears to mediate LPS stimulation
of cells that do not express mCD14 (10, 11), such as
endothelial, epithelial and smooth-muscle
cells. CD14 is found in both serum and urine (5).
The binding of LPS to CD14 requires an acute phase protein,
LPS-binding protein (LBP) (16).
The relationship of mCD14, CD14, LPS and LBP is complicated.
At low concentrations of LPS,
LBP is essential for the binding of LPS to CD14, but at high
concentrations, LBP may actually
inhibit binding of LPS to CD14. In addition, CD14 may
compete with mCD14 for LPS (17) and
may serve to help clear LPS (18). These four factors thus
appear to participate in a complex
feedback mechanism of immune regulation involving both
up-regulation and down-regulation
of the inflammatory process triggered by LPS. It is loss of
control of this mechanism that
appears to lead to septic shock. LPS-bound CD14 signals
production of inflammatory cytokines
and other inflammatory proteins, but the mechanism of signal
transduction is unclear. Since
a GPI anchor is not transmembrane, there presumably is
another transmembrane protein on
cells through which LPS-bound CD14 transmits a signal (19).
In addition to its well known role in gram-negative
infections, CD14 likely serves other
functions as well. It recognizes soluble peptidylglycan from
gram-positive cell walls (20), and it
has been reported to bind apoptotic cells and induce their
phagocytosis (21).
Entrez Gene IDs:
929 (Human); 12475 (Mouse); 60350 (Rat); 100037938 (Porcine)
Alternate Names:
CD14 antigen; CD14 molecule; CD14; monocyte differentiation antigen CD14; Myeloid cell-specific leucine-rich glycoprotein
