Category:Method/Sample/PL/Acidic - Revision history

Editor at 08:17, 5 November 2010

2010-11-05T08:17:51Z

← Older revision		Revision as of 17:17, 5 November 2010
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			{{Taguchi/Header}}
	=Acidic phospholipids （PA, LPA, PS, LPS, PI）=		=Acidic phospholipids （PA, LPA, PS, LPS, PI）=
	{{Twocolumn\|		{{Twocolumn\|

Dbmanager at 02:47, 27 October 2010

2010-10-27T02:47:28Z

← Older revision		Revision as of 11:47, 27 October 2010
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	# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースカラムにかける		# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースカラムにかける
	# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する		# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する
	# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, ~~PIとそれぞれのリゾ体~~)を溶出する		# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PI とそれぞれのリゾ体)を溶出する
	# 溶出液を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする		# 溶出液を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする
	}}		}}

Dbmanager at 02:46, 27 October 2010

2010-10-27T02:46:49Z

← Older revision		Revision as of 11:46, 27 October 2010
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	# PBS　2mL　を添加し、氷上で激しく撹拌する（３分）		# PBS　2mL　を添加し、氷上で激しく撹拌する（３分）
	# 遠心分離（2000g 5分）し、上層を分取する		# 遠心分離（2000g 5分）し、上層を分取する
	# ~~ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースにかける~~		# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースカラムにかける
	# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する		# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する
	# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PIとそれぞれのリゾ体)を溶出する		# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PIとそれぞれのリゾ体)を溶出する
	# 溶出液を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする		# 溶出液を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする
	}}		}}

Dbmanager at 08:29, 26 October 2010

2010-10-26T08:29:03Z

← Older revision		Revision as of 17:29, 26 October 2010
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	# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する		# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する
	# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PIとそれぞれのリゾ体)を溶出する		# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PIとそれぞれのリゾ体)を溶出する
	# ~~溶出液の溶媒を窒素気流で留去後、残存した水溶液約0~~.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする		# 溶出液を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする
	}}		}}

Dbmanager at 07:59, 26 October 2010

2010-10-26T07:59:17Z

← Older revision		Revision as of 16:59, 26 October 2010
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	# 細胞の1-ブタノール懸濁液　2mLを調製し、内標準　LPA(17:0), PA(14:0/14:0)など各50 pmolを添加する		# 細胞の1-ブタノール懸濁液　2mLを調製し、内標準　LPA(17:0), PA(14:0/14:0)など各50 pmolを添加する
	# PBS　2mL　を添加し、氷上で激しく撹拌する（３分）		# PBS　2mL　を添加し、氷上で激しく撹拌する（３分）
	# ~~遠心分離（2000rpm~~ 5分）し、上層を分取する		# 遠心分離（2000g 5分）し、上層を分取する
	# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースにかける		# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースにかける
	# ~~クロロホルム／メタノール~~(1:1) 3 x 1mLで洗浄する		# カラムに結合した中性脂質をクロロホルム／メタノール(1:1) 3 x 1mLで洗浄する
	# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, ~~and PI,それぞれのリゾ体~~)を溶出する		# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, PIとそれぞれのリゾ体)を溶出する
	# 溶出液の溶媒を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.~~1mLを混和する~~		# 溶出液の溶媒を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和し試料とする
	}}		}}

Dbmanager at 07:50, 26 October 2010

2010-10-26T07:50:21Z

← Older revision		Revision as of 16:50, 26 October 2010
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	\|		\|
	# 細胞の1-ブタノール懸濁液　2mLを調製し、内標準　LPA(17:0), PA(14:0/14:0)など各50 pmolを添加する		# 細胞の1-ブタノール懸濁液　2mLを調製し、内標準　LPA(17:0), PA(14:0/14:0)など各50 pmolを添加する
			# PBS　2mL　を添加し、氷上で激しく撹拌する（３分）
			# 遠心分離（2000rpm 5分）し、上層を分取する
			# ブタノール抽出液にメタノール1mLとクロロホルム1mLを混合したものを、DEAEセルロースにかける
			# クロロホルム／メタノール(1:1) 3 x 1mLで洗浄する
			# クロロホルム／メタノール／28%アンモニア水／氷酢酸 (200:100:10:6.7) 3 x 1mLで酸性脂質(PA, PS, and PI,それぞれのリゾ体)を溶出する
			# 溶出液の溶媒を窒素気流で留去後、残存した水溶液約0.05mLに水0.05mLと2-プロパノール0.1mLを混和する
	}}		}}

Dbmanager at 07:37, 26 October 2010

2010-10-26T07:37:22Z

← Older revision		Revision as of 16:37, 26 October 2010
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	# After removing organic solvent from each eluate under nitrogen gas, 0.05 mL of water and 0.1 mL of 2-propanol were added to about 0.05 mL of residue.		# After removing organic solvent from each eluate under nitrogen gas, 0.05 mL of water and 0.1 mL of 2-propanol were added to about 0.05 mL of residue.
	\|		\|
	# ~~日本語~~		# 細胞の1-ブタノール懸濁液　2mLを調製し、内標準　LPA(17:0), PA(14:0/14:0)など各50 pmolを添加する
	}}		}}

Dbmanager at 07:22, 26 October 2010

2010-10-26T07:22:56Z

← Older revision		Revision as of 16:22, 26 October 2010
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	=Acidic phospholipids （PA, LPA, PS, LPS, PI）=		=Acidic phospholipids （PA, LPA, PS, LPS, PI）=
	{{Twocolumn\|		{{Twocolumn\|
	# Cultured cells ~~(3 to 6 × 106 per 100 mm plate)~~ were scraped with 2 mL of ice-cold butanol, and 50 pmol of LPA(17:0) and PA(14:0/14:0) were added as an internal standard.		# Cultured cells were scraped with 2 mL of ice-cold butanol, and 50 pmol of LPA(17:0) and PA(14:0/14:0) were added as an internal standard.
	# 2 mL of ~~water~~ was added to ~~each~~ suspension, followed by vigorous shaking for 3 min while cooling with ice.		# 2 mL of PBS was added to suspension, followed by vigorous shaking for 3 min while cooling with ice.
	# Samples were centrifuged at ~~1000g~~ for 5 min, and the upper layer was collected.		# Samples were centrifuged at 2000g for 5 min, and the upper layer was collected.
	# After the addition of 1 mL of methanol and 1 mL of chloroform, each sample was applied to a DEAE-cellulose column.		# After the addition of 1 mL of methanol and 1 mL of chloroform, each sample was applied to a DEAE-cellulose column.
	# Column-bound lipids were washed with ~~chloroform (1 mL) and~~ chloroform/methanol (1:1) (3 × 1 mL) ~~in series~~.		# Column-bound lipids were washed with chloroform/methanol (1:1) (3 × 1 mL).
	# Acidic lipids, containing PA, PS, and PI, and their lyso forms, were eluted with chloroform/methanol/28% aqueous ammonia/acetic acid (200:100:10:6.7) (3× 1 mL).		# Acidic lipids, containing PA, PS, and PI, and their lyso forms, were eluted with chloroform/methanol/28% aqueous ammonia/acetic acid (200:100:10:6.7) (3× 1 mL).
	# After removing organic solvent from each eluate under nitrogen gas, 0.05 mL of water and 0.1 mL of 2-propanol were added to about 0.05 mL of residue.		# After removing organic solvent from each eluate under nitrogen gas, 0.05 mL of water and 0.1 mL of 2-propanol were added to about 0.05 mL of residue.

Editor: New page: =Acidic phospholipids （PA, LPA, PS, LPS, PI）= {{Twocolumn| # Cultured cells (3 to 6 × 106 per 100 mm plate) were scraped with 2 mL of ice-cold butanol, and 50 pmol of LPA(17:0) and PA...

2010-10-10T01:29:58Z

New page: =Acidic phospholipids （PA, LPA, PS, LPS, PI）= {{Twocolumn| # Cultured cells (3 to 6 × 106 per 100 mm plate) were scraped with 2 mL of ice-cold butanol, and 50 pmol of LPA(17:0) and PA...

New page

=Acidic phospholipids （PA, LPA, PS, LPS, PI）=
{{Twocolumn|
# Cultured cells (3 to 6 × 106 per 100 mm plate) were scraped with 2 mL of ice-cold butanol, and 50 pmol of LPA(17:0) and PA(14:0/14:0) were added as an internal standard.
# 2 mL of water was added to each suspension, followed by vigorous shaking for 3 min while cooling with ice.
# Samples were centrifuged at 1000g for 5 min, and the upper layer was collected.
# After the addition of 1 mL of methanol and 1 mL of chloroform, each sample was applied to a DEAE-cellulose column.
# Column-bound lipids were washed with chloroform (1 mL) and chloroform/methanol (1:1) (3 × 1 mL) in series.
# Acidic lipids, containing PA, PS, and PI, and their lyso forms, were eluted with chloroform/methanol/28% aqueous ammonia/acetic acid (200:100:10:6.7) (3× 1 mL).
# After removing organic solvent from each eluate under nitrogen gas, 0.05 mL of water and 0.1 mL of 2-propanol were added to about 0.05 mL of residue.
|
# 日本語
}}