McClean: Cycloheximide Stock Solution - Revision history

Megan N McClean: /* Notes */

2011-10-06T16:06:27Z

Notes

←Older revision		Revision as of 16:06, 6 October 2011
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	Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!		Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

-	'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when ~~weiging~~ it out and please make sure the balance area is clean.	+	'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weighing it out and please make sure the balance area is clean when you are finished.

	Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.		Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.

Megan N McClean at 16:05, 6 October 2011

2011-10-06T16:05:57Z

←Older revision		Revision as of 16:05, 6 October 2011
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	==Overview==		==Overview==
	This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you should check this concentration does what you think it does whenever you develop a new protocol using cycloheximide). Each person in the lab makes up their own stock of cycloheximide and keeps it in their own -20°C box.		This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you should check this concentration does what you think it does whenever you develop a new protocol using cycloheximide). Each person in the lab makes up their own stock of cycloheximide and keeps it in their own -20°C box.
-

	==Materials==		==Materials==

Megan N McClean: /* Materials */

2011-10-06T16:05:46Z

Materials

←Older revision		Revision as of 16:05, 6 October 2011
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	*Cycloheximide (MP Biomedicals, Cat #100183)		*Cycloheximide (MP Biomedicals, Cat #100183)
	*DMSO (Fluka 41639, Ultra for molecular biology; stored in the Flammables cabinet)		*DMSO (Fluka 41639, Ultra for molecular biology; stored in the Flammables cabinet)
-

	==Procedure==		==Procedure==

Megan N McClean: /* Notes */

2011-10-06T16:05:22Z

Notes

←Older revision		Revision as of 16:05, 6 October 2011
Line 17:		Line 17:

	'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weiging it out and please make sure the balance area is clean.		'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weiging it out and please make sure the balance area is clean.
		+
	Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.		Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.

Megan N McClean: /* Procedure */

2011-10-06T16:05:09Z

Procedure

←Older revision		Revision as of 16:05, 6 October 2011
Line 11:		Line 11:

	==Procedure==		==Procedure==
-	Weigh out the appropriate amount of cycloheximide and add it to the appropriate amount of DMSO to get a 50mg/ml solution. Use the Botstein lab's balance as it is much more accurate than ours. It seems to make the most sense to aim for about 100mg (but if it is slightly over or under that is fine) and then put this in a 2ml eppendorf or similar tube (you can use a 15ml conical if you are going to make a larger volume) and add the appropriate amount of DMSO to this based on what you weighed out.	+	Weigh out the appropriate amount of cycloheximide and add it to the appropriate amount of DMSO to get a 50mg/ml solution. Use the Botstein lab's balance as it is much more accurate than ours. It seems to make the most sense to aim for about 100mg (but if it is slightly over or under that is fine) and then put this in a 2ml eppendorf or similar tube (you can use a 15ml conical if you are going to make a larger volume) and add the appropriate amount of DMSO to this based on what you weighed out. Cycloheximide is bad for you (see below) so be careful with it!

	==Notes==		==Notes==

Megan N McClean: /* Notes */

2011-10-06T16:04:42Z

Notes

←Older revision		Revision as of 16:04, 6 October 2011
Line 15:		Line 15:
	==Notes==		==Notes==
	Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!		Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!
		+
	'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weiging it out and please make sure the balance area is clean.		'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weiging it out and please make sure the balance area is clean.
	Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.		Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.

Megan N McClean: /* Notes */

2011-10-06T16:04:29Z

Notes

←Older revision		Revision as of 16:04, 6 October 2011
Line 15:		Line 15:
	==Notes==		==Notes==
	Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!		Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!
-		+	'''*[[User:Megan N McClean\|Megan N McClean]]''': Cycloheximide is BAD for you. Check out the MSDS here: http://www.mpbio.com/product_info.php?family_key=02100183 or in the lab binder. Wear appropriate protective equipment (gloves, lab coat, glasses) when weiging it out and please make sure the balance area is clean.
-	'''*[[User:Megan N McClean\|Megan N McClean]]''': ~~I generally find it useful to make a series of 1~~:~~10 dilutions of the α-factor (1mg~~/~~mL, 100μg~~/~~mL, 10μg~~/~~mL, 1μg/mL) when I first make~~ the ~~stock, as these are 1000x the concentrations I generally start out with for most experiments (so that I can add 1μL of stock solution per ml of media)~~. ~~I usually make 900μL of all of these dilute stocks~~ (ie, ~~start with 100μL of 1mg/ml stock +900μL of DMSO for the 100μg/mL stock~~, ~~from that take 100μL and add~~ it ~~to another 900μL of DMSO for the 10μg/mL,~~ and ~~then once more for the 1μg/mL stock). I then divide the 900μL into 100μL aliquots so that I don't have to freeze/thaw~~ the ~~entire stock too often~~.	+
-		+
	Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.		Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.

Megan N McClean: /* Procedure */

2011-10-06T16:01:43Z

Procedure

←Older revision		Revision as of 16:01, 6 October 2011
Line 11:		Line 11:

	==Procedure==		==Procedure==
-	~~Shake~~ the ~~new vial~~ of ~~α-factor~~ to ~~get all of~~ the ~~power on the bottom~~ of ~~the vial~~. ~~Remove~~ the ~~lid and pipet 1ml of DMSO into the vial of α-factor from Sigma~~. ~~Pipet up~~ and ~~down~~ and ~~put~~ the ~~lid back on an swirl to dissolve all~~ of ~~the alpha-factor into the~~ DMSO~~. Then remove the liquid and aliquot it in 100μL eppendorfs into individual eppendorfs and store these in your own -20°C box~~.	+	Weigh out the appropriate amount of cycloheximide and add it to the appropriate amount of DMSO to get a 50mg/ml solution. Use the Botstein lab's balance as it is much more accurate than ours. It seems to make the most sense to aim for about 100mg (but if it is slightly over or under that is fine) and then put this in a 2ml eppendorf or similar tube (you can use a 15ml conical if you are going to make a larger volume) and add the appropriate amount of DMSO to this based on what you weighed out.

	==Notes==		==Notes==

Megan N McClean: New page: ==Overview== This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you ...

2011-10-06T15:55:56Z

New page:  ==Overview== This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you ...

New page

==Overview==
This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you should check this concentration does what you think it does whenever you develop a new protocol using cycloheximide). Each person in the lab makes up their own stock of cycloheximide and keeps it in their own -20°C box.

==Materials==
*Cycloheximide (MP Biomedicals, Cat #100183)
*DMSO (Fluka 41639, Ultra for molecular biology; stored in the Flammables cabinet)

==Procedure==
Shake the new vial of α-factor to get all of the power on the bottom of the vial. Remove the lid and pipet 1ml of DMSO into the vial of α-factor from Sigma. Pipet up and down and put the lid back on an swirl to dissolve all of the alpha-factor into the DMSO. Then remove the liquid and aliquot it in 100μL eppendorfs into individual eppendorfs and store these in your own -20°C box.

==Notes==
Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

'''*[[User:Megan N McClean|Megan N McClean]]''': I generally find it useful to make a series of 1:10 dilutions of the α-factor (1mg/mL, 100μg/mL, 10μg/mL, 1μg/mL) when I first make the stock, as these are 1000x the concentrations I generally start out with for most experiments (so that I can add 1μL of stock solution per ml of media). I usually make 900μL of all of these dilute stocks (ie, start with 100μL of 1mg/ml stock +900μL of DMSO for the 100μg/mL stock, from that take 100μL and add it to another 900μL of DMSO for the 10μg/mL, and then once more for the 1μg/mL stock). I then divide the 900μL into 100μL aliquots so that I don't have to freeze/thaw the entire stock too often.

Please sign your name to your note by adding <font face="courier"><nowiki>'''*~~~~''':</nowiki></font> to the beginning of your tip.

==Contact==
*'''[[User:Megan N McClean|Megan N McClean]] 14:01, 6 October 2011 (EDT)'''

or instead, [[Talk:{{PAGENAME}}|discuss this protocol]].



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