Alzheimer’s disease is a major global concern. Developing a cure or therapy is especially difficult since the disease progresses slowly and irreversibly over a decade or two. Diagnosis is especially challenging, since the symptoms are common to a plurality of causes.
Shimadzu Scientific (Columbia, MD) recently introduced a screening service to assay blood plasma for the presence of biomarkers amyloid peptides for Alzheimer’s disease (AD) at the low-pM level. This service can be used to identify and stratify patients before they are symptomatic. Presymptomatic studies are believed to be essential for understanding the etiology of AD and offer the best hope of arresting progression to dementia. The service is for research use only (RUO) since the assay has not yet met the FDA’s requirements for clinical diagnostics.
Experimental
The authors describe a simple immunoprecipitation protocol followed by matrix assisted-laser desorption ionization (MALDI-TOF) reading of the plate. The initial patient cohort was 62 normal, 30 with mild cognitive impairment, and 29 diagnosed with AD. Patient age ranged from 60 to 90 years. Later, blood samples were collected from a cohort of Australians. Blinded tests were performed independently by Shimadzu in Japan.
The supplemental material for the published paper provides a detailed description of the protocol used in Japan.1 For immune precipitation of the amyloid from plasma, BioLegend (San Diego, CA) clone 6E10 was conjugated to DynaBeads M-270 (Thermo Fisher Scientific, Waltham, MA). Isotopically labeled Aβ1-38 peptide (AnaSpec, San Jose, CA) was used as a common internal standard at 10-pM concentration for all sample peptides. After 1-hour incubation and washing, the peptides were taken up in 70% acetonitrile (+5 mM HCl) and transferred to wells on the MALDI plate.
MALDI plate
The MALDI plate (900 µm, Hudson Surface Technology, Ft. Lee, NJ) was prespotted with α-cyano-4-hydroxycinnaminic acid and methane diphosphonic acid.
Mass spectrometer
Mass spectra were recorded on an AXIMA Performance MALDI-linear TOF MS (Shimadzu) using a 337-nm nitrogen laser.
Data system
Mass++ V2.0 software (Shimadzu) provided the peak intensities and m/z values. Each peptide had a unique MS pattern. Aβ1-42 and Aβ1-40/Aβ1-42 ratio along with the amyloid precursor protein (APP)669-711 appeared to have the highest correlation with AD. Coefficients of variation were generally within 2–8%. Further, the results were confirmed by positron emission tomography (PET), which was selected as the standard of truth. Accuracy is about 90%.
Careful study of this immunoprecipitation/MS-TOF protocol did not reveal any obvious difficult experimental procedures or unique instrumental capabilities.
Shimadzu’s brochure provides a concise summary:
“This study was the first successful blinded validation of blood-based biomarkers that used two independent large datasets from two different countries. Their report not only demonstrated the high accuracy of the blood test, but also the reliability and reproducibility. The findings show that Shimadzu’s Amyloid MS Service can be used to detect early signs of Alzheimer’s disease in people with no obvious symptoms such as memory loss.”
It will be interesting to see how Shimadzu selects and executes a business model to provide these results in a clinical setting.
Reference
- Nakamura, A.; Kaneko, N. et al. High performance plasma amyloid-β biomarkers for Alzheimer’s disease. Nature Feb 8, 2018, 554, 249 & ff.; doi:10.1038/nature 25456.
Robert L. Stevenson, Ph.D., is Editor Emeritus, American Laboratory/Labcompare; e-mail: [email protected]