Automated microparticle enzyme immunoassay for neural thread protein in cerebrospinal fluid from Alzheimer's disease patients.

An automated microparticle enzyme immunoassay (MEIA) with the IMx analyzer for the detection of neural thread protein (NTP) in cerebrospinal fluid (CSF) from Alzheimer's disease (AD) patients was developed. This assay uses monoclonal antibodies produced against the purified pancreatic form of the protein. The assay employs one monoclonal antibody covalently coupled to the microparticle to capture immunoreactive material in CSF or brain tissue. The second monoclonal antibody was conjugated to alkaline phosphatase and serves as detection antibody. The assay provides results in approximately 45 minutes with a sensitivity of 60 pg/ml (3 fmoles/ml). The titration curve of both normal and AD CSF resulted in a linear relationship with respect to the volume of CSF used. A similar relationship was observed when normal and AD brain tissue extracts were serially diluted. The molecular weight of NTP in CSF was approximately 20 kD as determined by gel filtration method under non-denaturing conditions. The recovery for pancreatic thread protein (PTP) spiked in either normal or AD CSF was 104% and 108%, respectively. Intra-, inter-, and total assay CVs (coefficient of variation) for controls were less than 2.9%, 3.3% and 3.0%, respectively. This assay will provide a useful tool in the study of the Alzheimer's disease and may help research in diagnosis and prognosis of Alzheimer's disease and related disorders.

A sandwich enzyme immunoassay for detecting and measuring Alzheimer's disease--associated proteins in human brain tissue.

A simple, fast, and reliable immunoassay has been developed to detect and measure Alzheimer's disease-associated proteins (ADAPs) in human brain tissue. This assay, called ALZ-EIA (brain), was developed for Abbott's quantum system, in which 1/4-in. beads are used as solid phase. The beads were coated with polyclonal IgG purified from sera obtained from rabbits immunized with a highly enriched Alzheimer's disease brain protein fraction for ADAP. The antigen (ADAP) is effectively captured by the polyclonal IgG coated on the beads. ALZ-50 (a mouse monoclonal IgM) is used as the detection antibody, and bound ALZ-50 is subsequently quantified by a horseradish peroxidase-linked goat antimouse IgM and appropriate substrate. The assay is linear up to 0.5 absorbance units (r = 0.9), reproducible (CV less than 10%), sensitive, and specific. With preformulated reagents and standard supplies, the assay is simple and rapid; 120 data points can readily be generated in about 4 hr.