Mouse tumors were homogenized by stainless steel beads in 1% RIPA buffer (0.5% sodium deoxycholate, 50 mmol/L Tris-base, pH 7.4, 150 mmol/L NaCl, 1% Triton X-100 and 0.1% SDS). ELISA array was performed using the mouse IFN-γ (RK00019, ABclonal, China), mouse IL-2 (RK00007, ABclonal, China) and mouse TNF-α (RK00027, ABclonal, China) ELISA Kit according to the manufacturer's instructions.
Rk00007
Manufactured by ABclonal
Sourced in China
RK00007 is a lab equipment product. It is a device used for the centrifugation of samples in a laboratory setting. The core function of this product is to separate components of a mixture based on their differential rates of sedimentation within a centrifugal field.
Automatically generated - may contain errors
2 protocols using rk00007
1
Cytokine Profiling of Mouse Tumors
2
Quantification of Cytokine Levels in Mouse Serum
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For the quantitative determination of cytokine levels in serum, the blood of mice was collected from the orbital veins, then serum was separated at room temperature and stored at -80°C. The concentrations of interleukin-2 (IL-2) (pg/mL) (RK00007, ABclonal, China), IL-4 (pg/mL) (RK00036, ABclonal, China), and IL-10 (pg/mL) (RK00016, ABclonal, China) were determined using commercial enzyme-linked immunosorbent assay (ELISA) kits.
Binding ELISA was conducted to verify the affinity of antibody in the supernatant from pVAX-α-PD-1-transfected cells. The procedure carried out was as follows: 96-well plates were pre-coated with 10 μg/mL of PD-1 protein (Sigma) overnight at 4°C. They were subsequently washed with 0.1% Tween 20/PBS and then blocked with 5% bovine serum albumin (BSA) in a humid chamber for 1 hour at room temperature. After blocking, plates were washed three times and incubated at room temperature with supernatant from cells for 1 hour. Next, plates were washed and incubated with HRP goat anti-mouse lgG (H+L) secondary antibody at a dilution of 1:10,000 for 1 hour. The optical density was measured at 450 nm with an automatic ELISA reader (Synergy H1 Microplate Reader, BioTek, USA).
Binding ELISA was conducted to verify the affinity of antibody in the supernatant from pVAX-α-PD-1-transfected cells. The procedure carried out was as follows: 96-well plates were pre-coated with 10 μg/mL of PD-1 protein (Sigma) overnight at 4°C. They were subsequently washed with 0.1% Tween 20/PBS and then blocked with 5% bovine serum albumin (BSA) in a humid chamber for 1 hour at room temperature. After blocking, plates were washed three times and incubated at room temperature with supernatant from cells for 1 hour. Next, plates were washed and incubated with HRP goat anti-mouse lgG (H+L) secondary antibody at a dilution of 1:10,000 for 1 hour. The optical density was measured at 450 nm with an automatic ELISA reader (Synergy H1 Microplate Reader, BioTek, USA).
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