Lipofectamine Plasmid Transfection - Revision history

Irith at 21:00, 11 July 2012

2012-07-11T21:00:28Z

← Older revision		Revision as of 21:00, 11 July 2012
Line 21:		Line 21:
	#Add DNA/Lipofectamine complexes (~500 uL) to the 2 mL of media on the cells		#Add DNA/Lipofectamine complexes (~500 uL) to the 2 mL of media on the cells
	#Gently rock plate to mix		#Gently rock plate to mix
	#After ~4h re-feed cells with normal media.		#After ~4h aspirate media and re-feed cells with normal media. (It is usually possible to leave overnight).

	Protocol adapted from [http://tools.invitrogen.com/content/sfs/manuals/lipofectamine2000_man.pdf Product Manual]		Protocol adapted from [http://tools.invitrogen.com/content/sfs/manuals/lipofectamine2000_man.pdf Product Manual]

Davebrid at 14:18, 30 November 2009

2009-11-30T14:18:38Z

← Older revision		Revision as of 14:18, 30 November 2009
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	*Cells in a 6 well dish, plated and at >90% confluence		*Cells in a 6 well dish, plated and at >90% confluence
	*Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)		*Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)

			==Required Amounts==
			*Calculate DNA to transfect
			Lipofectamine(in uL) = DNA(in ug) 2.5
			OptiMEM(in uL, equal for DNA and Lipofectamine) = Lipofectamine(in uL) 25

	==Protocol (6 well dish)==		==Protocol (6 well dish)==

Davebrid: /* Protocol (6 well dish) */

2009-08-13T14:45:22Z

Protocol (6 well dish)

← Older revision		Revision as of 14:45, 13 August 2009
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	==Protocol (6 well dish)==		==Protocol (6 well dish)==
	#Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics.		#Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics (DMEM with 10% FBS, no PSG).
	#For each well prepare:		#For each well prepare:
	##250uL OptiMEM plus 4 ug of DNA.		##250uL OptiMEM plus 4 ug of DNA.

Davebrid: /* Protocol (6 well dish) */

2009-08-12T18:18:30Z

Protocol (6 well dish)

← Older revision		Revision as of 18:18, 12 August 2009
Line 7:		Line 7:

	==Protocol (6 well dish)==		==Protocol (6 well dish)==
	#Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics~~, or alternatively change media before transfection to OptiMEM~~		#Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics.
	#For each well prepare:		#For each well prepare:
	##250uL OptiMEM plus 4 ug of DNA.		##250uL OptiMEM plus 4 ug of DNA.

Davebrid at 17:57, 12 August 2009

2009-08-12T17:57:57Z

← Older revision		Revision as of 17:57, 12 August 2009
Line 2:		Line 2:
	[[Category: Transfection]]		[[Category: Transfection]]

	Materials		==Materials==
	*Cells in a 6 well dish, plated and at >90% confluence		*Cells in a 6 well dish, plated and at >90% confluence
	*Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)		*Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)

			==Protocol (6 well dish)==
			#Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics, or alternatively change media before transfection to OptiMEM
			#For each well prepare:
			##250uL OptiMEM plus 4 ug of DNA.
			##250uL OptiMEM plus 10 uL of Lipofectamine.
			##Incubate ~5 minutes at room temperature.
			##Combine the two volumes of OptiMEM/DNA and OptiMEM/Lipofectamine.
			##Incubate ~20 min at room temperature.
			#Add DNA/Lipofectamine complexes (~500 uL) to the 2 mL of media on the cells
			#Gently rock plate to mix
			#After ~4h re-feed cells with normal media.

	Protocol adapted from [http://tools.invitrogen.com/content/sfs/manuals/lipofectamine2000_man.pdf Product Manual]		Protocol adapted from [http://tools.invitrogen.com/content/sfs/manuals/lipofectamine2000_man.pdf Product Manual]

Davebrid: intial page

2009-08-12T17:50:34Z

intial page

New page

[[Category: Cell Culture ]]
[[Category: Transfection]]

Materials
*Cells in a 6 well dish, plated and at >90% confluence
*Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)

Protocol adapted from [http://tools.invitrogen.com/content/sfs/manuals/lipofectamine2000_man.pdf Product Manual]