Protein Extraction and Immunoblotting Protocol

Protein samples from supernatants and cell lysates were prepared as previously described [25 (link)]. The membrane transfer parameters for pro-IL-1β, pro-caspase-1, caspase-1 p20, NLRP3, β-actin and His-tag were 200 mA/90 min. The primary antibody dilution rates were 1:1000 for targets of interleukin-1β (IL-1β; R&D Systems, Minneapolis, MN, USA), caspase-1 (p20) (Adipogen, Switzerland), and NLRP3 (Adipogen SA, Epalinges, Switzerland) and 1:5000 for targets of His-tag (Amylet Scientific, Wuhan, China) and β-actin (Proteintech, Wuhan, China).

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Zhao P., Li J., Li X., Dong J., Wang X., Zhang N., Li S., Sun M., Zhang X., Wang Z., Liang M., Li Y., Cao L, & Gong P. (2023). The NLRP3 inflammasome recognizes alpha-2 and alpha-7.3 giardins and decreases the pathogenicity of Giardia duodenalis in mice. Parasites & Vectors, 16, 85.

Publication 2023

IL-1β β-actin

Actin Antibody Caspase 1 Cell Dilution Il 1β Interleukin 1β Membrane Pro caspase 1 Protein

Corresponding Organization : Jilin Academy of Agricultural Sciences

Other organizations : Jilin University

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Variable analysis

independent variables

Protein samples from supernatants and cell lysates

dependent variables

Pro-IL-1β
Pro-caspase-1
Caspase-1 p20
NLRP3
β-actin
His-tag

control variables

Membrane transfer parameters for pro-IL-1β, pro-caspase-1, caspase-1 p20, NLRP3, β-actin and His-tag (200 mA/90 min)
Primary antibody dilution rates for interleukin-1β (IL-1β; 1:1000), caspase-1 (p20) (1:1000), NLRP3 (1:1000), His-tag (1:5000), and β-actin (1:5000)

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