Gasdermin-D (GSDMD) is a member of the gasdermin (Gsdm) protein family, and its cleavage by inflammatory cysteine proteases (caspases, CASPs) is a critical event in cell pyroptosis. The role and functions of GSDMD on mice and humans are widely studied, but its expression, structure, and function in other species are less known. In the present work, rabbit anti-porcine GSDMD (pGSDMD) polyclonal antibody was prepared by immunizing New Zealand white rabbits with prokaryotic expressed recombinant pGSDMD (rpGSDMD). The prepared polyclonal antibody showed good specificity in Western blot and indirect immunofluorescence (IIF) assays. Western blot results showed that the polyclonal antibody could recognize overexpressed pGSDMD in human embryonic kidney cells (HEK293T) and endogenously expressed pGSDMD in cultured intestinal porcine enterocytes (IPEC-J2) and porcine kidney cells (PK-15). Western blot also revealed that pGSDMD was expressed in the heart, liver, lung, kidney, gallbladder, and jejunum of pigs. HEK293T cells overexpressing GSDMD showed green fluorescence in the IIF assay only after being treated with 0.3% Triton-X 100, which indicated that the full-length pGSDMD was located in the plasma but not on the cell membrane. This work provides a useful tool and basic information for further studies on pGSDMD.

Gasdermin-D (GSDMD) 是 gasdermin (Gsdm) 蛋白家族的成员，其被炎性半胱氨酸蛋白酶 (caspases, CASPs) 裂解是细胞焦亡的关键事件。GSDMD 在小鼠和人类中的作用和功能已被广泛研究，但其在其他物种中的表达、结构和功能却鲜为人知。本工作以原核表达重组pGSDMD（rpGSDMD）免疫新西兰大白兔，制备兔抗猪GSDMD（pGSDMD）多克隆抗体。制备的多克隆抗体在蛋白质印迹和间接免疫荧光 (IIF) 测定中显示出良好的特异性。Western blot结果显示，该多克隆抗体可识别人胚肾细胞（HEK293T）中过表达的pGSDMD，并在培养的猪肠细胞（IPEC-J2）和猪肾细胞（PK-15）中内源性表达pGSDMD。Western印迹还显示pGSDMD在猪的心脏、肝脏、肺、肾脏、胆囊和空肠中表达。仅在用 0.3% Triton-X 100 处理后，过表达 GSDMD 的 HEK293T 细胞在 IIF 测定中显示绿色荧光，这表明全长 pGSDMD 位于血浆中，而不是位于细胞膜上。这项工作为进一步研究 pGSDMD 提供了有用的工具和基本信息。仅在用 0.3% Triton-X 100 处理后，过表达 GSDMD 的 HEK293T 细胞在 IIF 测定中显示绿色荧光，这表明全长 pGSDMD 位于血浆中，而不是位于细胞膜上。这项工作为进一步研究 pGSDMD 提供了有用的工具和基本信息。仅在用 0.3% Triton-X 100 处理后，过表达 GSDMD 的 HEK293T 细胞在 IIF 测定中显示绿色荧光，这表明全长 pGSDMD 位于血浆中，而不是位于细胞膜上。这项工作为进一步研究 pGSDMD 提供了有用的工具和基本信息。