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01000caa a22002652c 4500 |
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NLM087941325 |
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DE-627 |
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20250130182033.0 |
| 007 |
tu |
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231222s1996 xx ||||| 00| ||eng c |
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2 |
|a pubmed25n0293.xml
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|a (DE-627)NLM087941325
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| 035 |
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|a (NLM)8825159
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| 040 |
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|a DE-627
|b ger
|c DE-627
|e rakwb
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| 041 |
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|a eng
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| 100 |
1 |
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|a Voss, E W
|c Jr
|e verfasserin
|4 aut
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| 245 |
1 |
0 |
|a Detection of protease activity using a fluorescence-enhancement globular substrate
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| 264 |
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1 |
|c 1996
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| 336 |
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|a Text
|b txt
|2 rdacontent
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| 337 |
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|a ohne Hilfsmittel zu benutzen
|b n
|2 rdamedia
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| 338 |
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|a Band
|b nc
|2 rdacarrier
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| 500 |
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|a Date Completed 03.12.1996
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| 500 |
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|a Date Revised 28.09.2018
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| 500 |
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|a published: Print
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| 500 |
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|a Citation Status MEDLINE
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| 520 |
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|a Bovine serum albumin (BSA) highly derivatized with fluorescein isothiocyanate (FITC, isomer I) served as a fluorescent enhancement substrate to measure protease activity. In the native globular BSA structure, the fluorescence of the lysine-conjugated fluorescein moieties was quenched 98%. Proteolytic digestion of highly derivatized BSA with Pronase resulted in fluorescence enhancement of 4300%. Both alpha-chymotrypsin and proteinase K yielded lower but similar fluorescence enhancement values of 2880% and 2800%, respectively. Digestion of the fluorescein-BSA substrate with trypsin, which required basic amino acids for activity, showed fluorescence enhancement of 1480% reflecting the fluorescein-lysine thiocarbamyl linkage. When derivatized substrate was pretreated with a thiol-reducing agent prior to incubation with proteases, a relatively small increase in fluorescence was noted relative to the untreated substrate except in the case of Pronase. The minimum sensitivity of proteolytic activity, based on a comparison of untreated and reduced FITC25BSA was 32 x 10(-6) units for I ng proteinase K, 1 x 10(-3) units for 1 ng alpha-chymotrypsin and 10 x 10(-3) units for Pronase and trypsin (1 ng each). The fluorescence enhancement assay was suited for sensitive intensity measurements or as an endpoint assay to detect protease activity
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| 650 |
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4 |
|a Journal Article
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| 650 |
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7 |
|a Alkylating Agents
|2 NLM
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| 650 |
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7 |
|a Fluorescent Dyes
|2 NLM
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| 650 |
|
7 |
|a Iodoacetates
|2 NLM
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| 650 |
|
7 |
|a Serum Albumin
|2 NLM
|
| 650 |
|
7 |
|a Dithioerythritol
|2 NLM
|
| 650 |
|
7 |
|a 6892-68-8
|2 NLM
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| 650 |
|
7 |
|a Endopeptidases
|2 NLM
|
| 650 |
|
7 |
|a EC 3.4.-
|2 NLM
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| 650 |
|
7 |
|a Chymotrypsin
|2 NLM
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| 650 |
|
7 |
|a EC 3.4.21.1
|2 NLM
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| 650 |
|
7 |
|a Trypsin
|2 NLM
|
| 650 |
|
7 |
|a EC 3.4.21.4
|2 NLM
|
| 650 |
|
7 |
|a Endopeptidase K
|2 NLM
|
| 650 |
|
7 |
|a EC 3.4.21.64
|2 NLM
|
| 650 |
|
7 |
|a Pronase
|2 NLM
|
| 650 |
|
7 |
|a EC 3.4.24.-
|2 NLM
|
| 650 |
|
7 |
|a Fluorescein-5-isothiocyanate
|2 NLM
|
| 650 |
|
7 |
|a I223NX31W9
|2 NLM
|
| 700 |
1 |
|
|a Workman, C J
|e verfasserin
|4 aut
|
| 700 |
1 |
|
|a Mummert, M E
|e verfasserin
|4 aut
|
| 773 |
0 |
8 |
|i Enthalten in
|t BioTechniques
|d 1991
|g 20(1996), 2 vom: 29. Feb., Seite 286-91
|w (DE-627)NLM012627046
|x 1940-9818
|7 nnas
|
| 773 |
1 |
8 |
|g volume:20
|g year:1996
|g number:2
|g day:29
|g month:02
|g pages:286-91
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| 912 |
|
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|a GBV_USEFLAG_A
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| 912 |
|
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|a SYSFLAG_A
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| 912 |
|
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|a GBV_NLM
|
| 912 |
|
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|a GBV_ILN_21
|
| 912 |
|
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|a GBV_ILN_22
|
| 912 |
|
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|a GBV_ILN_39
|
| 912 |
|
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|a GBV_ILN_40
|
| 912 |
|
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|a GBV_ILN_50
|
| 912 |
|
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|a GBV_ILN_60
|
| 912 |
|
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|a GBV_ILN_62
|
| 912 |
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|a GBV_ILN_65
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| 912 |
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|a GBV_ILN_70
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| 912 |
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|a GBV_ILN_99
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| 912 |
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|a GBV_ILN_121
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| 912 |
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|a GBV_ILN_130
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| 912 |
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|a GBV_ILN_227
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| 912 |
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|a GBV_ILN_350
|
| 912 |
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|a GBV_ILN_618
|
| 912 |
|
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|a GBV_ILN_640
|
| 912 |
|
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|a GBV_ILN_754
|
| 912 |
|
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|a GBV_ILN_2001
|
| 912 |
|
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|a GBV_ILN_2002
|
| 912 |
|
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|a GBV_ILN_2003
|
| 912 |
|
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|a GBV_ILN_2005
|
| 912 |
|
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|a GBV_ILN_2006
|
| 912 |
|
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|a GBV_ILN_2007
|
| 912 |
|
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|a GBV_ILN_2008
|
| 912 |
|
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|a GBV_ILN_2009
|
| 912 |
|
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|a GBV_ILN_2010
|
| 912 |
|
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|a GBV_ILN_2012
|
| 912 |
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|a GBV_ILN_2015
|
| 912 |
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|a GBV_ILN_2018
|
| 912 |
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|a GBV_ILN_2023
|
| 912 |
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|a GBV_ILN_2035
|
| 912 |
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|a GBV_ILN_2040
|
| 912 |
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|a GBV_ILN_2060
|
| 912 |
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|a GBV_ILN_2099
|
| 912 |
|
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|a GBV_ILN_2105
|
| 912 |
|
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|a GBV_ILN_2121
|
| 912 |
|
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|a GBV_ILN_2470
|
| 951 |
|
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|a AR
|
| 952 |
|
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|d 20
|j 1996
|e 2
|b 29
|c 02
|h 286-91
|