|Detection of Human and Mouse Phospho-APP/Protease Nexin II (T668) by Western Blot. Western blot shows lysates of L‑929 mouse fibroblast cell line and HeLa human cervical epithelial carcinoma cell line untreated (-) or treated (+) with 1 μg/mL nocodazole for 16 hours. PVDF membrane was probed with 0.5 µg/mL of Rabbit Anti-Human/Mouse Phospho-APP/Protease Nexin II (T668) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2508), followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.|
|Phospho-APP/Protease Nexin II (T668) in Human Brain. APP/Protease Nexin II phosphorylated at T668 was detected in immersion fixed paraffin-embedded sections of human Alzheimer's brain (cortex) using Rabbit Anti-Human/Mouse Phospho-APP/Protease Nexin II (T668) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2508) at 10 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Rabbit HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS005) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.|
Amyloid precursor protein (APP) is a type I membrane protein with several human isoforms due to alternative splicing. APP-770, -751, and -733 contain a Kunitz protease inhibitor (KPI) domain (residue 291‑342) and APP-695 does not. APP is a cell surface molecule with many functions. It can be processed proteolytically in two different pathways. In one pathway, beta - and gamma -secretase cleave at the beta site between residue 670 and 671 and the gamma site between residue 711 and 714 to produce beta ‑amyloid peptide (A beta 40 and A beta 42), a major component in plaques found in brains of patients with Alzheimer's disease (1). The other pathway involves alpha -secretase that cleaves residues between 687 and 688. It is antiamyloidogenic due to its benign character and the prevention of the A beta peptide formation (2). Soluble APP containing the KPI domain, also referred to as protease nexin II, is a potent inhibitor of serine proteases and may have additional functions. For example, it may regulate the contact face of blood coagulation and limit thrombosis specially in brain due to its localization and coagulation factor XI inhibiting activity (3, 4).
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