Detection of endogenous B-type natriuretic peptide at very low concentrations in patients with heart failure.
BACKGROUND
The myocardium secretes B-type natriuretic peptide (BNP) in response to stimuli associated with heart failure (HF). However, high immunoreactive-BNP levels in patients with HF are associated with a paradoxical lack of natriuretic response. We hypothesized that commercially available assays for immunoreactive BNP do not reflect the bioactivity of the natriuretic peptide system, because they measure both unprocessed inactive pro-BNP and mature BNP 1-32. We describe an assay for the detection of bioactive BNP 1-32 and confirm very low concentrations in plasma from HF patients.


RESULTS
We developed a quantitative mass spectrometry immunoassay to capture endogenous BNP peptides using high affinity antibodies. Bound BNP and its truncated fragments were detected by matrix assisted laser desorption ionization-time of flight mass spectrometry based on their predicted masses. Mass spectrometry immunoassay revealed rapid in vitro degradation of BNP 1-32 in plasma, which requires plasma collection in the presence of high protease inhibitor concentrations. In 11 of 12 HF patients BNP 1-32 was detectable, ranging from 25 to 43 pg/mL. Several degraded forms of BNP were also detected at similarly low levels. In contrast, parallel measurements of immunoreactive BNP using the Biosite assay ranged from 900 to 5000 pg/mL.


CONCLUSIONS
Detection of endogenous BNP 1-32 requires special preservation of plasma samples. Mass spectrometry immunoassay technology demonstrates that HF patients have low levels of BNP 1-32. Commercially available immunoreactive-BNP assays overrepresent biological activity of the natriuretic peptide system because they cannot distinguish between active and inactive forms. This observation may, in part, explain the "natriuretic paradox."
Niederkofler
Eric E
EE
Scios Inc., Mountain View, CA, USA.
Kiernan
Urban A
UA
O'Rear
Jessica
J
Menon
Santosh
S
Saghir
Syed
S
Protter
Andrew A
AA
Nelson
Randall W
RW
Schellenberger
Ute
U
D000368:Aged; D000369:Aged, 80 and over; D001793:Blood Preservation / Q000379:methods; D004305:Dose-Response Relationship, Drug; D005260:Female; D006333:Heart Failure / Q000097:blood*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008875:Middle Aged; D020097:Natriuretic Peptide, Brain / Q000097:blood* / Q000187:drug effects / Q000378:metabolism; D009994:Osmolar Concentration; D011480:Protease Inhibitors / Q000008:administration & dosage
D011480:Protease Inhibitors; D020097:Natriuretic Peptide, Brain