Direct immunochemiluminescent assay for proBNP and total BNP in human plasma proBNP and total BNP levels in normal and heart failure.
BACKGROUND
Recent studies have shown that in addition to brain (or B-type) natriuretic peptide (BNP) and the N-terminal proBNP fragment, levels of intact proBNP are also increased in heart failure. Moreover, present BNP immunoassays also measure proBNP, as the anti-BNP antibody cross-reacts with proBNP. It is important to know the exact levels of proBNP in heart failure, because elevation of the low-activity proBNP may be associated with the development of heart failure.


RESULTS
We therefore established a two-step immunochemiluminescent assay for total BNP (BNP+proBNP) and proBNP using monoclonal antibodies and glycosylated proBNP as a standard. The assay enables measurement of plasma total BNP and proBNP within only 7 h, without prior extraction of the plasma. The detection limit was 0.4 pmol/L for a 50-µl plasma sample. Within-run CVs ranged from 5.2%-8.0% in proBNP assay and from 7.0%-8.4% in total BNP assay, and between-run CVs ranged from 5.3-7.4% in proBNP assay and from 2.9%-9.5% in total BNP assay, respectively. The dilution curves for plasma samples showed good linearity (correlation coefficients   = 0.998-1.00), and analytical recovery was 90-101%. The mean total BNP and proBNP in plasma from 116 healthy subjects were 1.4 ± 1.2 pM and 1.0 ± 0.7 pM, respectively, and were 80 ± 129 pM and 42 ± 70 pM in 32 heart failure patients. Plasma proBNP levels significantly correlate with age in normal subjects.


CONCLUSIONS
Our immunochemiluminescent assay is sufficiently rapid and precise for routine determination of total BNP and proBNP in human plasma.
Department of Medicine and Clinical Science, Kyoto University Graduate School of Medicine, Kyoto, Japan. nishikim@kuhp.kyoto-u.ac.jp
Toshio
T
Nishikimi
Hiroyuki
H
Okamoto
Masahiro
M
Nakamura
Naoko
N
Ogawa
Kazukiyo
K
Horii
Kiyoshi
K
Nagata
Yasuaki
Y
Nakagawa
Hideyuki
H
Kinoshita
Chinatsu
C
Yamada
Kazuhiro
K
Nakao
Takeya
T
Minami
Yoshihiro
Y
Kuwabara
Koichiro
K
Kuwahara
Izuru
I
Masuda
Kenji
K
Kangawa
Naoto
N
Minamino
Kazuwa
K
Nakao
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D002138:Calibration; D016022:Case-Control Studies; D003429:Cross Reactions; D042241:Early Diagnosis; D005260:Female; D006333:Heart Failure / Q000097:blood* / Q000175:diagnosis; D006801:Humans; D007118:Immunoassay / Q000592:standards*; D057230:Limit of Detection; D008163:Luminescent Measurements; D008297:Male; D008875:Middle Aged; D020097:Natriuretic Peptide, Brain / Q000097:blood*; D015588:Observer Variation; D011498:Protein Precursors / Q000097:blood*; D012015:Reference Standards; D015203:Reproducibility of Results
D000911:Antibodies, Monoclonal; D011498:Protein Precursors; D020097:Natriuretic Peptide, Brain