| 1 | Define the biological question first | Match the biomarker to amyloid pathology, tau pathology, neurodegeneration, or disease-related symptoms. | Amyloid-β, phosphorylated tau, total tau, neurofilament light, structural or metabolic imaging. | Cerebrospinal fluid, blood, magnetic resonance imaging, or positron emission tomography. | Different biological processes and stages of Alzheimer disease. | No single biomarker captures the full disease process; multimodal designs are often more informative. |
| 2 | Prioritize biomarkers of amyloid pathology | Identify evidence of cerebral amyloid accumulation, a defining feature of Alzheimer disease biology. | Cerebrospinal fluid Aβ42, Aβ40, and the Aβ42/Aβ40 ratio; amyloid positron emission tomography. | Lumbar puncture or amyloid-sensitive brain imaging. | Presence or absence of amyloid plaques in the brain. | Cerebrospinal fluid measures and imaging findings require method-specific interpretation and validated thresholds. |
| 3 | Include biomarkers of tau pathology | Assess disease-related tau phosphorylation and neurofibrillary tangle burden. | Plasma or cerebrospinal fluid phosphorylated tau, total tau, and tau positron emission tomography. | Blood, cerebrospinal fluid, or tau-sensitive brain imaging. | Biological evidence associated with Alzheimer-type tau pathology and disease progression. | Different phosphorylated tau forms are not interchangeable; assay platform and specimen handling affect results. |
| 4 | Add neurodegeneration and neuronal injury measures | Quantify downstream neuronal damage and overall disease-related injury. | Neurofilament light, total tau, hippocampal volume, cortical thickness, and fluorodeoxyglucose positron emission tomography. | Blood or cerebrospinal fluid; magnetic resonance imaging or metabolic imaging. | Neuronal injury, atrophy, or altered brain glucose metabolism. | These measures are generally less specific for Alzheimer disease and may also change in other neurological conditions. |
| 5 | Consider blood-based biomarkers for scalable screening | Support prescreening, longitudinal monitoring, and recruitment for confirmatory studies. | Plasma phosphorylated tau, plasma Aβ42/Aβ40, and neurofilament light. | Venous blood or dried blood spot specimens, depending on the validated method. | A less invasive indication of amyloid or tau-related biology and neuronal injury. | Pre-analytical controls, population diversity, kidney function, comorbidities, and assay validation are important. |
| 6 | Check analytical and clinical validation | Determine whether the assay produces reliable measurements and clinically meaningful classification. | Analytical precision, sensitivity, specificity, reproducibility, reference ranges, and concordance with established measures. | Validated laboratory assay, imaging protocol, or centralized testing workflow. | Confidence that results are technically consistent and relevant to the intended research use. | Results from different assays should not be compared directly without harmonization or cross-platform validation. |
| 7 | Plan for longitudinal and clinical-context data | Relate biomarker changes to cognition, functional status, treatment exposure, and disease stage. | Serial plasma or cerebrospinal fluid measures, repeated imaging, cognitive scores, and functional assessments. | Repeated visits using standardized collection, processing, and assessment procedures. | Disease trajectory, treatment response, progression risk, and the relationship between pathology and clinical symptoms. | Timing, attrition, practice effects, and changes in assay procedures can influence longitudinal interpretation. |