人胞內(nèi)細(xì)胞因子檢測刺激方案

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人細(xì)胞因子	細(xì)胞來源	活化-極化培養(yǎng)	培養(yǎng)時(shí)間	再次刺激	胞內(nèi)阻斷劑
G-CSF	PBMC	LPS (1 ug/mL)	24hr	-	Monensin
GM-CSF	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	5hr	-	Monensin
GIFN gamma	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	5hr	-	Brefeldin A
IL-1 alpha	PBMC	LPS (1 ug/mL)	24hr	-	Monensin
IL-1 beta	PBMC	LPS (100ng/mL)	4hr	-	Brefeldin A
IL-1RA	PBMC	LPS (100ng/mL)	24hr	-	Brefeldin A
IL-2	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	4-6hr	-	Brefeldin A
IL-4	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	4-6hr	-	Brefeldin A
IL-5	CD4	Th2 polarizing cultures	6d	PMA (50ng/mL) + Ionomycin (1ug/mL) (5hr)	Brefeldin A
IL-6	PBMC	LPS (100ng/mL)	24hr	-	Brefeldin A
IL-9	CD4	h2 polarizing culturesT	6d	PMA (50ng/mL) + Ionomycin (1ug/mL) (5hr)	Monensin
IL-10	CD4	Th2 polarizing cultures	6d	PMA (50ng/ml) + Ionomycin (1ug/ml) (5hr)	Monensin
IL-12/ IL-23 (p40)	PBMC	hIFN gamma (100ng/mL) (2hr)/LPS (100ng/mL) (22hr)	2hr/22hr	-	Brefeldin A
IL-13	CD4	anti-CD3 (10ug/mL, immobilized) + anti-CD28 (2ug/mL, soluble) + IL-2 (10ng/mL) + IL-4 (20ng/mL) (2d); IL-2 (10ng/mL) + IL-4 (20ng/mL) (3d)	2d/3d	PMA (5ng/ml) + Ionomycin (500ng/ml) (4hr)	Brefeldin A
IL-17A	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	5hr	-	Monensin
IL-21	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	4-7hr or 12-18hr	-	Brefeldin A
IL-22	CD4	Th17 polarizing cultures	6d	PMA (50ng/ml) + Ionomycin (1ug/ml) (5hr)	Brefeldin A
IL-23 p19	PBMC	hGM-CSF (40ng/mL) +hIL-4 (40ng/mL)	6d	LPS (1ug/mL) (24hr)	Monensin
MCP-1/ CCL2	PBMC	LPS (1ug/mL)	24hr	-	Monensin
RANTES/ CCL5	PBMC	LPS (1ug/mL)	24hr	-	Monensin
TNF alpha	PBMC	PMA (30-50ng/mL)/Iono (1ug/mL)	5hr	-	Brefeldin A
TNF beta	PBMC	Th1 polarizing cultures	6d	PMA (50ng/mL) + Ionomycin (1ug/mL) (5hr)	Monensin
注： PBMC=外周血單核細(xì)胞peripheral blood mononuclear cell；PMA=佛波酯Phorbol Myristate Acetate; calcium ionophore; 鈣離子載體 iono=離子霉素ionomycin; LPS=脂多糖Lipopolysaccharide; 4hr=4 hours culture; 2d=2 days culture. GolgiStop/GolgiPlug: 一種胞內(nèi)阻斷劑，可以用BFA/Monensin替代。來源：http://us.ebioscience.com/resources/best-protocols/flow-cytometry/flow-cytometry-intracellular-staining-quick-guide.htm

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