Bone marrow was isolated from two tibia and two femurs per mouse using 1 mL ice cold PBS. After light trituration, cells and debris were separated from extracellular fluid by centrifugation for 5 min at 400 × g. Fluid was aliquoted and frozen in liquid nitrogen. A Mouse CXCL12/SDF-1α Quantikine ELISA kit (R&D Systems) was used as per the manufacturer’s protocol. Bone marrow extracellular fluid preps were measured undiluted. Plates were read at 450 nm wavelength on a SpectraMAX190 and analyzed using the SoftMax Pro software (Molecular Devices, San Jose, CA).
Mouse cxcl12 sdf 1α quantikine elisa kit
Manufactured by R&D Systems
The Mouse CXCL12/SDF-1α Quantikine ELISA kit is a quantitative sandwich enzyme immunoassay designed to measure mouse CXCL12/SDF-1α levels in cell culture supernates, serum, and plasma. The kit utilizes a monoclonal antibody specific for mouse CXCL12/SDF-1α that has been pre-coated onto a microplate.
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3 protocols using mouse cxcl12 sdf 1α quantikine elisa kit
1
Quantifying CXCL12 in Mouse Bone Marrow
2
Measuring Cytokines and Chemokines in Bone Marrow
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For ELISA, BM supernatant was generated by flushing the BM plug from bones from one hind leg (femur + tibia) in 300 µl containing equal volumes of RayBio 2x Cell Lysis Buffer (Raybiotech/Lucerna Chem) and 2× complete ULTRA Protease Inhibitor Cocktail (Sigma-Aldrich). Levels of IFNα and IFNγ in serum and BM supernatants were tested using the VeriKine kit (PBL Assay Sciences) or Ready-SET-Go! Kit (eBioscience), respectively. SCF levels were determined using Mouse SCF ELISA (Raybiotech), and CXCL12 protein concentration was assayed with the Mouse CXCL12/SDF-1 α Quantikine ELISA Kit (R&D Systems) according to manufacturer’s instructions.
3
Temporal Dynamics of VEGF and SDF-1α in HLI
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Concentrations of vascular endothelial growth factor (VEGF) and stromal cell–derived factor‐1 (SDF‐1) α proteins in murine blood plasma samples were determined by ELISA kits (mouse VEGF ELISA kit, R&D Systems, and mouse CXCL12/SDF‐1α Quantikine ELISA kit, respectively), according to manufacturer's instructions.18 (link) Plasma levels of VEGF and SDF‐1α in sham operation and at day 7 after HLI surgery were measured every 6 hours (6:00, 12:00, 18:00, and 24:00) in both the control and jet‐lag groups.
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