Hypothesis: Cerebrospinal fluid (CSF) can be identified by using an enzyme-linked immunosorbent assay (ELISA) for Dickkopf-related protein 3 (DKK3).
Background: Cerebrospinal fluid leakage from the subarachnoid space is a potentially alarming condition that, left unrepaired, may result in increased risk of meningitis and encephalitis. Current biochemical methods of CSF leak detection involve using beta-2-transferrin-based or beta trace protein-based assays, both of which, at present, have limitations that hinder practical clinical application. This study presents the immunological detection of the CSF-enriched protein DKK3 as a method for detection of a CSF leak.
Methods: Antibodies against DKK3 were generated in rabbits and goats immunized with recombinant human DKK3. Varying dilutions and combinations of human CSF and serum were tested on immunoblots and sandwich ELISA using antibodies to DKK3.
Results: ELISA data show that there is a negligible amount of detectable DKK3 in serum samples compared with CSF samples. Inclusion of sera (up to 30%) in a sample containing CSF failed to produce a positive signal, whereas concentrations of CSF as low as 1% produced a positive signal. The minimum concentration required for reliable CSF detection in a sandwich ELISA was determined to be 0.5 μl.
Conclusion: ELISA sandwich assays for DKK3 can reliably detect the presence of as little as 0.5 μl of human CSF, even in the presence of excessive serum. This study provides quantitative evidence of the utility of DKK3 immunoreactivity as an assay for the presence of CSF in samples that contain contaminating sera. The robustness of this assay has allowed for the development of a rapid, point of care test for the detection of CSF in clinical and surgical setting.