CHARACTERIZATION OF NEW POLYCLONAL ANTIBODIES SPECIFIC FOR 40 AND 42 AMINO-ACID LONG AMYLOID-BETA PEPTIDES - THEIR USE TO EXAMINE THE CELL BIOLOGY OF PRESENILINS AND THE IMMUNOHISTOCHEMISTRY OF SPORADIC ALZHEIMERS-DISEASE AND CEREBRAL AMYLOID ANGIOPATHY CASES

Background: In Alzheimer's disease (AD), the main histological lesion is a proteinaceous deposit, the senile plaque, which is mainly compose d of a peptide called A beta. The aggregation process is thought to oc cur through enhanced concentration of A beta 40 or increased productio n of the more readily aggregating 42 amino acid-long A beta 42 species . Materials and Methods: Specificity of the antibodies was assessed by dot blot, Western blot, ELISA, and immunoprecipitation procedures on synthetic and endogenous A beta produced by secreted HK293 cells. A be ta and p3 production by wild-type and mutated presenilin l-expressing cells transiently transfected with beta APP751 was monitored after met abolic labeling and immunoprecipitation procedures. Immunohistochemica l analysis was performed on brains of sporadic and typical cerebrovasc ular amyloid angiopathy (CAA) cases. Results: Dot and Western blot ana lyses indicate that IgG-purified fractions of antisera recognize nativ e and denaturated A beta s. FCA3340 and FCA3542 display full specifici ty for A beta 40 and A beta 42, respectively. Antibodies immunoprecipi tate their respective synthetic A beta species but also A beta s and t heir related p3 counterparts endogenously secreted by transfected huma n kidney 293 cells. This allowed us to show that mutations on presenil in 1 triggered similar increased ratios of A beta 42 and its p342 coun terpart over total A beta and p3. ELISA assays allow detection of abou t 25-50 pg/ml of A beta s and remain linear up to 750 to 1500 pg/ml wi thout any cross-reactivity. FCA18 and FCA3542 label diffuse and mature plaques of a sporadic AD case whereas FCA3340 only reveals the mature lesions and particularly labels their central dense core. Ln a CAA ca se, FCA18 and FCA3340 reveal leptomeningeal and cortical arterioles wh ereas FCA3542 only faintly labels such structures. Conclusions: Polycl onal antibodies exclusively recognizing A beta 40 (FCA3340) or A beta 42 (FCA3542) were obtained. These demonstrated that FAD-linked preseni lins similarly affect both p342 and A beta 42, suggesting that these m utations misroute the beta APP to a compartment where gamma-secretase, but not alpha-secretase, cleavages are modified. Overall, these antib odies should prove useful for fundamental and diagnostic approaches, a s suggested by their usefulness for biochemical, cell biological, and immunohistochemical techniques.