Human cxcl12 sdf 1α quantikine elisa kit

The method used for the quantitative determination of CRP, IL-1β and SDF-1α was the enzyme-linked immunosorbent assay (ELISA) method. The protocol for each of the three ILs was performed following the instructions of R&D Systems Human Kit designated for each cytokine in part (Human CRP Quantikine ELISA Kit, IL-1β Human IL-1β/IL-1β F2 Quantikine ELISA Kit and Human CXCL12/SDF-1α Quantikine ELISA Kit, R&D Systems Inc., Minneapolis, MN). The normal range of values for CRP was between 1.09 and 4.291 mg/L, with a mean of 1.769 mg/L, with the limit of detection 0.005–0.022 ng/mL (mean value 0.010 ng/mL) and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was 3.8–4.3% and for inter-assay precision 6–7%. For IL-1β, the normal range of values was between 1 and 3.9 pg/mL, with the limit of detection 1 pg/mL and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was in the range 2.8–8.5% and for inter-assay precision, 4.1–8.4%. For SDF-1α, the normal range of values was between 1330 and 2720 pg/mL (mean 1830 pg/mL), with the limit of detection 1.0–47 pg/mL (mean value 18 pg/mL) and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was between 3.4% and 3.9% and for inter-assay precision, 8.2–13.4%

Huh7 TIMP-1 cells were grown for 48 h to approximately 90% confluence. Then, these cells were cultured with 0.5% FBS DMEM for 36 h. After centrifugation, the supernatants was harvested, passed through a sterile Millipore filter with a 0.45 mm polyvinylidenedifluoride membrane to generated conditioned medium from Huh7 TIMP-1 cells. The conditioned medium from Huh7 Vector cells was obtained in the same manner. All conditioned medium were maintained in liquid nitrogen for later use.
To detect the level of soluble SDF-1 secreted by LFs or CAFs, 2 × 10⁵ LFs or CAFs were seeded into 6-well plates and grown for 48 h, and the supernatant was harvested for ELISA. The human CXCL12/SDF-1α quantikine ELISA kit from R&D Systems (Minneapolis, MN, USA) was used to perform ELISAs for SDF-1 measurement according to the manufacturer's instructions. In the same manner, we examined the level of soluble TIMP-1 in conditioned medium from Huh7 TIMP-1 and Huh7 Vector cells. There were 3 replicates for each measurement, and the assessment was repeated six times.

Serum SDF-1α from breast cancer patients or similar-aged healthy women was quantified by ELISA using the Human CXCL12/SDF-1α Quantikine ELISA Kit (R&D Systems). Concentrations were calculated by comparing the sample absorbance to standard curves.

CXCL12 was measured on patients’ serum using the human CXCL12/SDF1α Quantikine ELISA kit (R&D Systems, Minneapolis, MN, USA), according to the manufacturer’s instructions. For each experiment, a titration curve was prepared using serial dilutions of the standard CXCL12 of the kit. The curve was then used to extrapolate the CXCL12 concentration in the samples.

The method used for the quantitative determination of CRP, IL-1β and SDF-1α was the enzyme-linked immunosorbent assay (ELISA) method. The protocol for each of the three ILs was performed following the instructions of R&D Systems Human Kit designated for each cytokine in part (Human CRP Quantikine ELISA Kit, IL-1β Human IL-1β/IL-1β F2 Quantikine ELISA Kit and Human CXCL12/SDF-1α Quantikine ELISA Kit, R&D Systems Inc., Minneapolis, MN). The normal range of values for CRP was between 1.09 and 4.291 mg/L, with a mean of 1.769 mg/L, with the limit of detection 0.005–0.022 ng/mL (mean value 0.010 ng/mL) and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was 3.8–4.3% and for inter-assay precision 6–7%. For IL-1β, the normal range of values was between 1 and 3.9 pg/mL, with the limit of detection 1 pg/mL and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was in the range 2.8–8.5% and for inter-assay precision, 4.1–8.4%. For SDF-1α, the normal range of values was between 1330 and 2720 pg/mL (mean 1830 pg/mL), with the limit of detection 1.0–47 pg/mL (mean value 18 pg/mL) and no significant cross-reactivity or interference; the coefficient of variance for intra-assay precision was between 3.4% and 3.9% and for inter-assay precision, 8.2–13.4%