PMID- 25933703
OWN - NLM
STAT- MEDLINE
DCOM- 20160301
LR  - 20171116
IS  - 1096-0309 (Electronic)
IS  - 0003-2697 (Linking)
VI  - 482
DP  - 2015 Aug 1
TI  - Fluorimetric screening assay for protein carbonyl evaluation in biological 
      samples.
PG  - 55-61
LID - S0003-2697(15)00198-0 [pii]
LID - 10.1016/j.ab.2015.04.021 [doi]
AB  - Many assays are available for the detection of protein carbonyls (PCs). 
      Currently, the measurement of PC groups after their derivatization with 
      2,4-dinitrophenol hydrazine (DNPH) is widely used for measuring protein oxidation 
      in biological samples. However, this method includes several washing steps. In 
      this context, we have developed a rapid, sensitive, and accurate fluorimetric 
      method adapted to 96-well microplates for the convenient assessment of protein 
      carbonyl level in biological samples. The method reported here is based on the 
      reaction of carbonyl content in proteins with 
      7-hydrazino-4-nitrobenzo-2,1,3-oxadiazole (NBDH) to form highly fluorescent 
      derivatives via hydrazone formation. PCs were determined using the DNPH and NBDH 
      assays in fully reduced bovine serum albumin (BSA) and plasma and liver 
      homogenates obtained from healthy control rats up the addition of various amounts 
      of HOCl-oxidized BSA (OxBSA). Using the NBDH assay, PC concentrations as low as 
      0.2 nmol/mg were detected with precision as low as 5%. Matrix-assisted laser 
      desorption/ionization time-of-flight (MALDI-TOF) mass spectroscopy was used to 
      successfully identify the formation of the NBDH adducts after derivatization with 
      standard oxidized peptides. Finally, the two methods were further used for PC 
      determination in plasma and liver samples from diabetic and normal rats, showing 
      that the NBDH assay can be reliably used in biological experiments.
CI  - Copyright (c) 2015 Elsevier Inc. All rights reserved.
FAU - Stocker, P
AU  - Stocker P
AD  - Aix Marseille Universite, CNRS, UMR 7273, ICR-SMBSO, 13397 Marseille Cedex 20, 
      France. Electronic address: p.stocker@univ-amu.fr.
FAU - Ricquebourg, E
AU  - Ricquebourg E
AD  - Aix Marseille Universite, CNRS, UMR 7273, ICR-SMBSO, 13397 Marseille Cedex 20, 
      France.
FAU - Vidal, N
AU  - Vidal N
AD  - Aix Marseille Universite, CNRS, UMR 7273, ICR-SMBSO, 13397 Marseille Cedex 20, 
      France; SARL YELEN, 13820 Ensues la Redonne, France.
FAU - Villard, C
AU  - Villard C
AD  - Centre de Recherches en Encologie Biologique et Onco-Pharmacologie (CRO2), 
      Aix-Marseille Universite, Institut National de la Sante et de la Recherche 
      Medicale (INSERM), 13385 Marseille, France.
FAU - Lafitte, D
AU  - Lafitte D
AD  - Centre de Recherches en Encologie Biologique et Onco-Pharmacologie (CRO2), 
      Aix-Marseille Universite, Institut National de la Sante et de la Recherche 
      Medicale (INSERM), 13385 Marseille, France.
FAU - Sellami, L
AU  - Sellami L
AD  - Centre de Recherches en Encologie Biologique et Onco-Pharmacologie (CRO2), 
      Aix-Marseille Universite, Institut National de la Sante et de la Recherche 
      Medicale (INSERM), 13385 Marseille, France.
FAU - Pietri, S
AU  - Pietri S
AD  - Aix Marseille Universite, CNRS, UMR 7273, ICR-SMBSO, 13397 Marseille Cedex 20, 
      France.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20150428
PL  - United States
TA  - Anal Biochem
JT  - Analytical biochemistry
JID - 0370535
RN  - 0 (7-nitrobenzoxadiazole)
RN  - 0 (Blood Proteins)
RN  - 0 (Dinitrophenols)
RN  - 0 (Hydrazines)
RN  - 0 (Oxadiazoles)
RN  - 27432CM55Q (Serum Albumin, Bovine)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Blood Proteins/chemistry/*metabolism
MH  - Cattle
MH  - Diabetes Mellitus, Experimental/metabolism
MH  - Dinitrophenols/chemistry
MH  - Fluorometry/*methods
MH  - Hydrazines/chemistry
MH  - Liver/chemistry/*metabolism
MH  - Oxadiazoles/chemistry
MH  - Oxidation-Reduction
MH  - *Protein Carbonylation
MH  - Rats
MH  - Serum Albumin, Bovine/chemistry
MH  - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
OTO - NOTNLM
OT  - Fluorescence
OT  - MALDI-TOF
OT  - NBDH
OT  - Oxidized BSA
OT  - Protein carbonyls
EDAT- 2015/05/03 06:00
MHDA- 2016/03/02 06:00
CRDT- 2015/05/03 06:00
PHST- 2015/02/05 00:00 [received]
PHST- 2015/04/20 00:00 [revised]
PHST- 2015/04/22 00:00 [accepted]
PHST- 2015/05/03 06:00 [entrez]
PHST- 2015/05/03 06:00 [pubmed]
PHST- 2016/03/02 06:00 [medline]
AID - S0003-2697(15)00198-0 [pii]
AID - 10.1016/j.ab.2015.04.021 [doi]
PST - ppublish
SO  - Anal Biochem. 2015 Aug 1;482:55-61. doi: 10.1016/j.ab.2015.04.021. Epub 2015 Apr 
      28.