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No. 17 (PC 17)
Development of C-reactive protein assay for human saliva
Yumi Narita, Taku Shirakawa
Department of Biophysics, Graduate School of Health Sciences, Kobe University, Japan
BACKGROUND: Measurement of C-reactive protein (CRP) in saliva may be able to be used for the
early detection of the inflammatory disorder of infants or home care patients. However, the reports about
the correlation between salivary and serum CRP vary among researchers. Such differences might be
caused in the step of pretreatment or collection of saliva. In this study, we examined these steps in order
to develop a high-sensitivity assay for salivary CRP.
METHODS: Salivary CRP concentration was determined by sandwich enzyme-linked immunoassay
(ELISA). To improve the sensitivity of the assay, Ampliflu TM Red (Sigma) was used as a fluorogenic
substrate. Saliva was collected by drooling and/or saliva collection swab. Saliva samples were diluted
1:1 in sample diluent containing Tween20 and/or cation, then centrifuge at 1500g for 5 minutes.
Concentrations of Tween20 and cations were examined in 0 to 1% and 0 to 1000mM, respectively. The
validity of ELISA was examined by intra-assay precision and recovery test. Saliva and serum samples
from 28 healthy volunteers were collected, and then CRP concentrations were measured in order to
evaluate the correlation of salivary and serum CRP.
RESULTS: 500mmol/L MgCl2 and 0.25%Tween20 were best at enhancing the CRP fluorescent signal.
Both MgCl2 and Tween20 were essential to detect strong signal. Under these conditions intra-assay CV
was 3.0%, and recovery was 102-107%. CRP concentration decreased 30% in samples collected with
saliva collection swabs in comparison with drooling. CRP was measurable in saliva samples (84.8 to
4104.3pg/mL) and serum samples (26.0 to 2015.4ng/mL). There was a positive correlation between
salivary and serum CRP (r=0.77).
CONCLUSIONS: Strong enhancement of CRP fluorescent signal was observed in the presence of
MgCl2 and Tween20. Salivary CRP concentration was affected by collection method. Salivary CRP and
serum CRP moderately correlated. This result will support for the validation of salivary CRP as an
alternative marker of inflammation.
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