Tau151-391 tagged with hemagglutinin (HA) was overexpressed in HEK-293FT cells. The cells, 48 h after transfection, were lysed in phosphate-buffered saline (PBS) containing 50 mM NaF, 1 mM Na3VO4, 1 mM AEBSF, 5 mM benzamidine, and 10 μg/ml each of aprotinin, leupeptin, and pepstatin by probe sonication at 20% power for 2 min. The cell lysates were centrifuged for 10 min at 10,000×g. The extract containing HA-tau151-391 was stored at − 80 °C until use.

Various amounts of AD tau fractions were dotted on nitrocellulose membrane and dried at 37 °C for 1 h. The membrane was blocked with 5% fat-free milk-TBS for 1 h and incubated with the above cell extract containing HA-tau151-391 overnight. After washing with TBST, captured HA-tau151-391 was detected by incubating with anti-HA in 5% milk-TBST and processed as described above for immuno-dot blots.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.