Standard Curve
Example of Human CXCL9 standard curve in Assay Diluent TA2. Plotted are the background-subtracted data.
This standard curve is provided for demonstration only.
Product Details
Product Details
Product Specification
| Antigen | CXCL9 |
| Immunogen | Recombinant Protein |
| Antibody Type | Recombinant mAb |
| Reactivity | Hu |
| Purification | Protein A |
| Stability & Storage | 12 months from date of receipt / reconstitution, 2 to 8°C as supplied. |
Kit
| Precision | Intra-assay: 2.6% Inter-assay: 4.9% |
| Sample type | Cell culture supernatant; Plasma |
| Assay type | Sandwich (quantitative) |
| Sensitivity | 5.04 pg/mL |
| Range | 31.25 pg/mL – 2000 pg/mL |
| Recovery | Cell culture supernatant: 93% Heparin Plasma: 88% EDTA Plasma: 99% |
| Assay time | 60 minutes |
| Species reactivity | Hu |
Background
Human CXCL9, also known as Monokine Induced by Interferon-Gamma (MIG), is a small cytokine belonging to the CXC chemokine family that plays a critical role in immune responses. It is primarily secreted by monocytes, macrophages, and endothelial cells in response to stimulation by interferon-gamma (IFN-γ), with its expression further enhanced by TNF. As an ELR-negative chemokine, CXCL9 selectively attracts activated T lymphocytes and Th1 cells by binding to its receptor, CXCR3, without affecting neutrophils. The protein functions as a potent inhibitor of angiogenesis and contributes to anti-tumor immunity by promoting the infiltration of effector T cells into the tumor microenvironment. The mature, secreted human CXCL9 protein consists of 103 amino acids and has a molecular weight of approximately 11.7 kDa. It is synthesized as a 125-amino-acid precursor with a 22-residue signal peptide that is cleaved to yield the active form. Due to its central role in Th1-mediated inflammation and its involvement in conditions ranging from autoimmune diseases to cancer, CXCL9 is an important research target and a potential biomarker for various pathological states.
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ELISA
Spike Recovery
The recovery of Human CXCL9 was evaluated in activated samples spiked with concentrations spanning the entire assay range.
Spike-and-dilution Linearity
To evaluate assay linearity, three samples were spiked with high levels of Human CXCL9 in different matrices and serially diluted with the corresponding Calibrator Diluent to fall within the assay's dynamic range.
Dilution linearity
The concentrations of Human CXCL9 were measured and interpolated from the target standard curves and corrected for sample dilution.
The leftmost bar in the graph represents neat biological samples. "Undiluted" was defined as the neat biological sample (100%). The subsequent bars represent two-fold serial dilutions of this reference. The mean target concentration was determined to be 69.8 pg/mL in Human EDTA plasma.
Dilution Linearity
The concentrations of Human CXCL9 were measured and interpolated from the target standard curves and corrected for sample dilution.
The leftmost bar in the graph shows biological samples diluted in Assay Diluent to 25% concentration. "undiluted" was defined as 25% biological sample. The subsequent bars represent two-fold serial dilutions of this reference. The mean target concentration was determined to be 198.3 pg/mL in Human heparin plasma.
Dilution Linearity
The concentrations of Human CXCL9 were measured and interpolated from the target standard curves and corrected for sample dilution.
The leftmost bar in the graph shows biological samples diluted in Assay Diluent to 20% concentration. "undiluted" was defined as 20% biological sample. The subsequent bars represent two-fold serial dilutions of this reference. The mean target concentration was determined to be 5228.4 pg/mL in Human peripheral blood mononuclear cells.
Determination of Minimum Detectable Dose (MDD)
The MDD was determined using three independent lots of assay components. For each lot, 19 replicate measurements of the diluent (zero calibrator) were performed. The mean (AVERAGE) and standard deviation (STDEV) of the 19 replicates were calculated. The MDD for each lot was then calculated according to the following formula:
MDD = 2 × STDEV + AVERAGE
Intra-Assay Precision (Precision within an assay)
Three samples of known concentration were tested fifteen times on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays)
Three samples of known concentration were tested in separate assays to assess inter-assay precision. Assays were performed with at least three lots of components.
HOOK Effect Threshold
The upper limit of the HOOK effect was established at 100× the highest calibrator concentration (equivalent to a 2-log10 increase). No HOOK effect was observed below this threshold, confirming that samples within this concentration range yield reliable quantitative results without signal depression.
Cross-reactivity
The cross-reactivity between Mouse CXCL9 and Human CXCL9 was assessed by testing both proteins in the same assay system. Serial dilutions of each protein were measured, and the dose-response curves were compared. No significant cross-reactivity was observed when Mouse CXCL9 was tested in the Human CXCL9 assay, as the signals remained within the background range of the assay.
Comparison of Standard Curves from Three Kits
Standard curves from Company A, Company B, and Starter Onestep were compared.
Protocol Diagram
