Human Anti-Aβ42 Recombinant Antibody (clone scFv59) (CAT#: FAMAB-0160-CN)

Figure 1 Optimization of scFv59 expression in the brain via rAAV serotype 1, 2, and 5 and theirpossible cytotoxicity in mice

Three serotypes of rAAV (1, 2, and 5) encoding scFv59 were injected into the right lateralventricle of C57BL/6 mice and, 3 months after injection, scFv59 expressed in the brain wasvisualized by immunohistochemistry using anti-FLAG M2 antibody (scFv59 contains theFLAG sequence as a marker). (a) PBS-injected mice were used as negative controls. Nostaining is observed in PBS-injected mice. (b) Widespread, symmetrical expression ofscFv59 is seen in both sides of the neocortex and the hippocampus in mice injected withrAAV1-CAscFv59. (c) Expression of scFv59 is limited mostly to a small part of the righthippocampus in mice subjected to rAAV2-CAscFv59 injection. (d) Widespread, symmetrical expression of scFv59 in mice injected with rAAV5-CAscFv59, whichresembles the expression pattern by rAAV1-CAscFv59. Levels of scFv59 expression inrAAV1-CAscFv59-injected mice are comparable to those in rAAV5-CAscFv59-injectedmice. To assess cytotoxicity and inflammation potentially associated with AAV-mediatedbrain expression of scFv59, brain sections were subjected to TUNEL assay (e–h) andimmunohistochemistry (i–l), respectively, using anti-CD11b antibody for activatedmicroglia. Apoptotic cells identified by TUNEL are seen in the brain injected with rAAV1-CAscFv59 (f) but not in the brains injected with PBS (e), rAAV2-CAscFv59 (g), andrAAV5-CAscFv59 (h). The inset in (f) is a higher magnification of the boxed area. ScatteredCD11b-positive cells are seen in the hippocampus of mice injected with rAAV1-CAscFv59(j) but not in mice injected with PBS (i), rAAV2-CAscFv59 (k), and rAAV5-CAscFv59 (l).The inset in (j) is a higher magnification of one of CD11b-positive microglia indicated byarrow heads. Scale bars 2.0 mm (a–d) and 500 μm (e–l).

Kou, J., Kim, H., Pattanayak, A., Song, M., Lim, J. E., Taguchi, H., ... & Fukuchi, K. I. (2011). Anti-amyloid-β single-chain antibody brain delivery via AAV reduces amyloid load but may increase cerebral hemorrhages in an Alzheimer's disease mouse model. Journal of Alzheimer's Disease, 27(1), 23-38.

Figure 2 Five months after rAAV5-CAscFv59 injection, CSF (A) and hippocampal homogenates (B) were prepared and subjected to 10–20% SDS PAGE followed by western blotting using anti-FLAG M2 antibody.

(A) scFv59 is identified as an approximately 30 kDa fragment in the CSF isolated from mice subjected to rAAV5-CAscFv59 injection. Lane 1: Molecular size markers; lane 2: Ten ng of purified scFv59 as a positive control; lane 3 Blank; lane 4: Ten μl of CSF from a PBS-injected mouse; and lane 5: Ten μl of CSF from a rAAV5-CAscFv59-injected mouse. (B) scFv59 is seen as 30–35 kDa fragments in the hippocampus of mice subjected to rAAV5-CAscFv59 injection. Lane 1: Molecular size markers; lane 2: Ten ng of purified scFv59 as a positive control; lane 3: Hippocampal homogenate from a PBS-injected mouse; and lane 4 and 5: Hippocampal homogenates from rAAV5-CAscFv59-injected mice.

Kou, J., Kim, H., Pattanayak, A., Song, M., Lim, J. E., Taguchi, H., ... & Fukuchi, K. I. (2011). Anti-amyloid-β single-chain antibody brain delivery via AAV reduces amyloid load but may increase cerebral hemorrhages in an Alzheimer's disease mouse model. Journal of Alzheimer's Disease, 27(1), 23-38.