For thymocyte proliferation assay by BrdU incorporation, the mice with indicated genotype were injected intraperitoneally with BrdU at the dosage of 100 mg/kg of body weight and analyzed 16 hours after injection. Cells were surface-stained, and then intracellular staining of BrdU was carried out using an FITC or APC BrdU Flow kit (catalog no. 559619 or 552598, BD Biosciences) following the manufacturer’s instruction. For detection of thymocyte apoptosis, cells were stained with indicated surface antibodies and then stained with anti–annexin V and 7-AAD using the PE annexin V Apoptosis Detection Kit (catalog no. 559763, BD Biosciences) following the manufacturer’s instruction.
Fitc or apc brdu flow kit
Manufactured by BD
The FITC or APC BrdU Flow kit is a laboratory tool used to detect and quantify cell proliferation. It provides a method for the measurement of bromodeoxyuridine (BrdU) incorporation into cellular DNA during the S-phase of the cell cycle. The kit includes reagents for labeling, fixing, and staining cells to allow for flow cytometric analysis.
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2 protocols using fitc or apc brdu flow kit
1
Thymocyte Proliferation and Apoptosis Assay
2
CFA-Induced Inflammation and LTβR Signaling
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CFA-induced inflammation was induced by i.p. injection of 200–400 μl of a 1:1 emulsion of DPBS and CFA (Sigma-Aldrich). Mice were analyzed at the stated time points after CFA injection. PolyI:C treatments were performed intravenously at 1.4 mg/Kg/d. LTβR signaling was blocked with 200 μg of LTβR-Ig consisting of the LTβR ectodomain fused with the Fc domain of a mouse IgG1 antibody specific for hen egg lysozyme (Hel-Ig). Control experiments were performed by treatment with 200 μg HEL-Ig. Inflammatory cytokines TNF-α, IFN-γ, and IL-1β were blocked with anti-TNF (#BE0058; Bio-X-Cell), anti-INF-γ (#BE0055; Bio-X-Cell), or anti-IL-1β (#BE0246; Bio-X-Cell) antibodies at 100 μg/mouse injected intravenously via the tail vein or retro-orbital sinus immediately before CFA injection. For LTβR-Ig treatment at steady state, mice were injected with 100 μg LTβR-Ig i.v. once a week for 3 wk and analyzed 3 wk after the first injection. In vivo BrdU incorporation was performed by BrdU injection i.v. (1 mg/mouse). Flow cytometry detection of incorporated BrdU was performed with FITC or APC BrdU Flow Kit (BD Biosciences) by following the manufacturer’s protocol.
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