ON-TARGETplus Human ITGB4 (3691) siRNA - SMARTpool

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	siRNA concentration-30nM Replace medium 1h before transfection. Add diluted siRNA to siLentFect Reagent to make a mixture and incubate the mixture for 20 min at room temperature. Replace culture medium after 4h of transfection and incubate the plate at 37°C for 24 to 72 h.

Protocol tips
siRNA concentration-30nM Replace medium 1h before transfection. Add diluted siRNA to siLentFect Reagent to make a mixture and incubate the mixture for 20 min at room temperature. Replace culture medium after 4h of transfection and incubate the plate at 37°C for 24 to 72 h.

Publication protocol

DU145 human prostate cancer cells were obtained from ATCC (HTB-81) and maintained in high glucose DMEM with 10% fetal bovine serum (FBS), 1% glutamine, penicillin and streptomycin (GPS), and 1% sodium pyruvate (Invitrogen, Carlsbad, CA). Phase contrast microscopy was performed using a TE2000 microscope (Nikon) and RT SPOT camera with SPOT Advanced v4.0.9. software (Diagnostic Instruments, Inc., Sterling Heights, MI). Cells were transfected with siRNA using SilentFect (Biorad) in Opti-MEM I Reduced Serum Medium (Invitrogen), incubated for 4 hours, media changed, and cells used for assays at 48-72 hr. siRNAs were obtained from Thermo Scientific: ON-TARGETplus non-targeting control siRNA pool (D-001818-10-05), ON-TARGETplus human EPCAM siRNA pool (L-004568-01-0005), ON-TARGETplus human PLAU siRNA (L-006000-00-0005), ON-TARGETplus human ITGB4 siRNA pool (L-008011-00-0005). EPCAM and ITGB4 siRNAs were used at 30nM and PLAU siRNA used at 90nM for effective knockdown without toxicity.

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Manufacturer protocol

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