Approach to Selected Diseases: Alzheimer Disease (AD)
The hallmark pathological features of Alzheimer Disease (AD) in humans are amyloid plaques and neurofibrillary tangles. Over the past decade, an increasing number of genetically engineered mouse models for AD research have been developed to exhibit classic Alzheimer features. The timeline for development of disease features ranges from a few months to ~18 months. NSA has processed hundreds of human AD tissues and thousands of brains from AD mouse models.
NSA typically receives one hemisphere of each mouse brain for AD neurohistology while the other hemisphere is often held by the client for enzyme-linked immunosorbent assay (ELISA) analysis. NSA can process up to 40 mouse brain hemispheres coronally or 20 mouse brain hemispheres sagittally in each MultiBrain® Block (or 25 if the area of interest is the cerebrum). Alternatively, when both hemispheres are made available to NSA, 20 or 25 whole mouse brains can be processed sagittally in each block. The preference of the researcher determines which alignment is chosen.
With human tissue, NSA is capable of processing human brain and other large format sections, as in the image below, providing a unique opportunity to assess large contiguous cross-sections of tissue. More often however, NSA is asked to process multiple smaller samples from one or more brains using MultiBrain® Technology.
The standard NSA practice of encasing the brain tissue in gelatin provides a significant aid in the handling of tissue sections resulting in an improved final product.
Primary AD Endpoints: Amyloid Plaques and Neurofibrillary Tangles
The hallmark features of AD are amyloid plaques and neurofibrillary tangles. Amyloid-laden plaques exist as a diffuse form and as a more dense/mature form (congophilic). Amyloid is also found as deposits in vessels. Neurofibrillary tangles are present in cell bodies and in the neuropil as “neuropil threads.”
The choice of which stain(s) to use is based on the specific needs of the study. The Campbell-Switzer Alzheimer pathology stain, developed by NSA, is capable of showing all of the hallmark features of Alzheimer pathology, while other stains/antibody methods are capable of revealing more specific features. Please note that the Campbell-Switzer stain does not work on monkey tissue.
Strategic Approach to Amyloid Detection: Different approaches yield unique features
At specific times during an R&D cycle, different approaches to amyloid detection may be most useful. Each of the methods described has its own strengths and specialized advantage:
Campbell-Switzer Method: Dr. Bob Switzer and his former associate, Shannon Campbell, developed the Campbell-Switzer Alzheimer Pathology stain in the early 1980’s. Over the years, this unique stain has become a “work horse” tool for researchers, particularly in the earlier phases of R&D. Diffuse plaques are stained black and the more dense fibrillar amyloid forms are stained amber. Neurofibrillary tangles typically stain black.
Immunohistochemistry for Amyloid Plaque Detection: The antibody methods are intended to be very specific and reveal amyloid peptides that have been cleaved at a specific location. On the basis of a particular mouse model, treatment approach and other factors, a specific antibody can be more useful than others in revealing a specific measure of efficacy.
Some examples of antibody methods include:
|Abeta Amyloid (plaques)||Tau (neurofibrillary tangles)|
|Aß (1-40)||AT8 (pSer202+Thr205)|
|Aß (1-42)||Tau 46 (Epitope 404-441)|
|Aß (1-28)||Anti-Human Tau HT7 (Epitope 158-163)|
|4G8||Anti-Human Tau (Epitope 243-441)|
|6E10||phospho Tau Thr181|
|phospho Tau S396|
|phospho Tau S422|
Congo Red: Congo Red specifically reveals the more dense, fibrillar amyloid plaques (hence, congophilic plaques). When isolating these types of plaques as an endpoint, Congo Red is an effective solution, as is Thioflavin S.
Thioflavin S: As with Congo Red, Thioflavin S specifically reveals the more dense, fibrillar amyloid plaques (congophilic plaques), but with a fluorescent signal. This is a useful option if double fluorescent labeling is desired.
Alzheimer Endpoint: Inflammation
Inflammation occurs in conjunction with the hallmark pathologic features of AD and can be detected by examining microglia (Iba1 IHC) or astrocytes (GFAP IHC).
Alzheimer Mouse Model Plaque Chemoarchitecture
AD Endpoint: Disintegrative Debris
Images taken from sections of the brain of an old, >12 months, PS-1/APP mouse
AD Endpoint: Increase in Iron
Endpoints Overview and Table of Stains for AD
This table depicts the most commonly used stains for detecting various categories of markers used in Alzheimers research.
|Stain||Primary Pathology: Plaques and Tangles||Inflammation||Other Feature-Specific Endpoints|
|Aß 1-40 IHC||√|
|Aß 1-42 IHC||√|
|Aß 1-43 IHC||√|
|Aß 1-28 IHC||√|
|H & E||√||√||√|
|CD 11b IHC||√|
|Tau pSer 422||√||√|
|Anti-Human Tau HT7||√||√|