... wells, aspirate the media from the wells. Gently wash the cells with 1 mL of PBS and aspirate wash. Filter...added until 48–72 h after transduction. Gently aspirate the media from the cells. Add 1.5 mL of DMEM complete...colonies should be visible. Every 3–4 days, gently aspirate the media and replace it with fresh DMEM complete...

...Vortex primers, probe and master mix for 15 sec then spin 10 sec in a mini centrifuge and place on ice. Wipe...Volume 20 µL Vortex the master mix for 15 sec and spin in a mini centrifuge for 10 sec before use. Place...generated, use a 20–200 µL multichannel pipet to aspirate 40 µL of droplets. Pro-Tip To ensure that the ...45° angle. Count to 20 while slowly and gently aspirating the droplets. Transfer the droplets to a prechilled...

...each T-175 flask: Aspirate culture media and rinse once with 10 mL of PBS. Aspirate PBS and add 2 mL of...at ~80% confluence. Aspirate culture media and rinse once with 60 mL PBS. Aspirate PBS and add 35 mL of...

...Untransduced 0 1498.5 1.5 Mix the dilutions well Gently aspirate media from the cells. Add 1.5 mL of a viral dilution...calculating the titer. Incubate for 48–72 h. Gently aspirate media and replace with 1 mL of PBS. Calculate ...

... hood. Clean any spills immediately, and decontaminate as necessary. For large spills that cannot be easily...

... the cells at 37 °C, 5% CO 2 for ~20 h. Gently aspirate media, add 10 mL fresh DMEM complete containing...following morning. The following morning, carefully aspirate the media. Replace the media with 15 mL of DMEM...

...the media and chloroquine and mix well. Gently aspirate the media out of the previously seeded 10 cm plate...following morning. The following morning, carefully aspirate the media. Replace the media with 10 mL of DMEM...

...sample. Add 2 µL 10X reducing agent to each sample. Spin the sample briefly in the microcentrifuge. Heat ...the samples for 10 min at 100 °C in a heat block. Spin the sample briefly in the microcentrifuge. Remove...

... tissue/cell lysate. Pro-Tip Having your samples spin at 4°C helps reduce RNA degradation. If you don’...

...100 μg/mL ampicillin or carbenicillin. Day 2: 2. Spin down the cultures and use a miniprep kit to obtain...media from cells and pool with media from Day 4. Spin media at 1,250 rpm for 5 minutes to pellet any HEK...

...Repeat for each sample. Notes: Keep in mind that despite the accuracy of the NanoDrop, if two consecutive...

...transfer it into your container. Make sure that any spills produced during the protocol are cleaned up. Discard...

...Section 2: Fixing and permeabilizing cells Gently aspirate the media from the 24-well plate. Wash each well...

...isopropanol to precipitate the plasmid DNA. Either spin to pellet the DNA or apply the solution to a column...

...Incubate the cells with the virus for 48–72 h. Gently aspirate the media from the cells. Add 1.5 mL of DMEM complete...

... bit more than 200 mL (~220 mL) just in case we spill anything or have small errors in measurement. You...