Validation experiment designed to validate causal mechanisms targeting TREM2 in Mouse (5xFAD/TREM2-/-). Measures changes in biological function and validates hypothesis predictions.
Negative control: Wild-type age-matched littermates (no amyloid expected)
Antibody control: Sections stained with secondary antibody only (no primary)
Technical replicates: 3 IHC runs per antibody
Statistical Analysis
Compare % amyloid area between genotypes using two-tailed Student's t-test
Two-way ANOVA for age × genotype interaction
Significance threshold: p < 0.05 (Bonferroni corrected for multiple comparisons)
Effect size (Cohen's d) reported for all significant comparisons
Expected Outcomes
Expected Outcomes
Primary Outcomes
Increased amyloid burden in TREM2-/- mice: TREM2 KO mice are expected to show 30–60% greater plaque coverage vs TREM2+/+ 5xFAD mice at 6 months based on published literature (Wang et al. 2015; Jay et al. 2015)
Increased plaque number and size: TREM2 KO expected to increase diffuse/immature plaque count; compaction (Thioflavin-S) may be reduced
Accelerated temporal progression: Differences expected to be larger at 9 months than 6 months
...
Expected Outcomes
Primary Outcomes
Increased amyloid burden in TREM2-/- mice: TREM2 KO mice are expected to show 30–60% greater plaque coverage vs TREM2+/+ 5xFAD mice at 6 months based on published literature (Wang et al. 2015; Jay et al. 2015)
Increased plaque number and size: TREM2 KO expected to increase diffuse/immature plaque count; compaction (Thioflavin-S) may be reduced
Accelerated temporal progression: Differences expected to be larger at 9 months than 6 months
Secondary Outcomes
TREM2 KO mice may show increased soluble Aβ42/Aβ40 ratio in brain lysates
Reduced microglial clustering around plaques in TREM2-/- (assessed by IBA1 co-staining)
No gross phenotypic differences between groups expected
Negative/Null Result Interpretation
If no difference observed, may indicate TREM2 role is pathway-specific or age-dependent
Null result at 6 months should prompt re-analysis at 9+ months
Success Criteria
Success Criteria
Primary
[ ] ≥30% increase in cortical Aβ % area coverage in 5xFAD/TREM2-/- vs 5xFAD/TREM2+/+ (p < 0.05)
[ ] ≥30% increase in hippocampal Aβ % area coverage (p < 0.05)
[ ] Effect reproduces in both 6-month and 9-month cohorts
Secondary
[ ] Thioflavin-S+ plaque density quantified in both groups
[ ] IBA1 microglial density around plaques quantified