...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...solution through a 0.22 μm membrane. Aliquot 500–1000 μL into sterile tubes. Store the tubes at -20 °C...PEI such that the ratio of μg DNA:μg PEI is 1:3 (1000 μL total per 10 cm dish). Using transfer plasmid...lots of FBS can promote or inhibit transfection. Test a variety of brands and lots of FBS to find one ... plate in DMEM Complete in 10 cm tissue culture plates. Incubate the cells at 37 °C, 5% CO 2 for ~20 h...the table below for a possible range of ratios to test: Ratio of DNA:PEI μg of DNA μL of 1 mg/mL PEI 1:...Centrifuge the viral supernatant at 2100 rcf for 5 minutes to pellet any packaging cells that were collected...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Hemocytometer or cell ..., Nalgene, 565-0010 (or for larger volumes 1000 mL 0.45 µm Rapid-Flow PES Filtration Unit, Nalgene 167...solution through a 0.22 μm membrane. Aliquot 500–1000 μL into sterile tubes. Store the tubes at -80 °C... deionized water and adjust to a final volume of 1000 mL. Stir at RT until fully dissolved. Pro-Tip This...lots of FBS can promote or inhibit transfection. Test a variety of brands and lots of FBS to find one ...inactivated in the lab by heating to 56 °C for 30 minutes. 0.45 μm polyethersulfone (PES) filter system, ...and add 35 mL of 0.05% Trypsin/EDTA. Wait ~2-3 minutes for cells to detach. Gently tap the sides of the...

...mL conical collection tube. Centrifuge column at 1000 x g for 2 min to remove storage solution. Place ...container. Add 5 mL PBS to the column. Centrifuge at 1000 x g for 2 min to remove PBS. Repeat Steps 30–31 ...resin bed to fully absorb the sample. Centrifuge at 1000 x g for 2 min to collect the sample. Discard column...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment...tubes, VWR 76289-498 Before Starting Wipe down all pipettes and equipment with 10% bleach prior to use. Warm... Adjust pH to 7.0 Autoclave or filter sterilize 1000X protease inhibitor cocktail 25 mg leupeptin 50 mg...Spectrophotometer using the A280 IgG setting. Combine all eluates with measurable protein. Determine the protein ...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette CO 2 incubator Pipet controller...96-well plate, seed one of the corner wells with ~1000 cells. This will help you focus the microscope later... incubating the cells. Pro-Tip Use the well with 1000 cells to focus the microscope, then you can easily...PES filter or centrifugation at >500 x g for 5 minutes. Do not use the medium if the cells are overly ...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...solution through a 0.22 µm membrane. Aliquot 500–1000 µL into sterile tubes. Store the tubes at -80 °C...lots of FBS can promote or inhibit transfection. Test a variety of brands and lots of FBS to find one ... plate in DMEM complete in 10 cm tissue culture plates. Incubate the cells at 37 °C, 5% CO 2 for ~20 h...the table below for a possible range of ratios to test: Ratio of DNA:PEI Amount of DNA (μg) Volume of 1...

...filter, VWR 431229 0.22 µm PES filtering system, 1000 mL, VWR 431098 0.45 µm PES filtering system, 500... aliquots and store at -20 °C until use. BCD TFX 1000 mL BalanCD HEK293 Media 10 mL 10% Pluronic-F68 40...Biosafety Cabinet 4 °C Refrigerator Micropipettes and tips Pipettor and pipettes Benchtop centrifuge compatible...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment...Reagent Preparation instructions. Wipe down all pipettes and equipment with 10% bleach prior to use. Reagent...suggest preparing fresh solutions after one month. 1000X protease inhibitor cocktail 25 mg Leupeptin 50 mg...

...tips, 1000 µL, VWR 76322-154 Pipettes, 5 mL, VWR 89130-896 Pipettes, 10 mL, VWR 89130-898 Pipettes, 25 ...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Pipette controller Microcentrifuge...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment...25 mL, VWR 89130-900 Pipettes, 50 mL, VWR 89130-902 Gel loading tips, Corning 4853 Microcentrifuge tubes...mobility of the bands (sample AR0016 in Figure 1) indicates that the samples may not have been processed correctly...

... pipette 200–1000 µL single channel pipette Pipette controller Pipette tips and pipettes Spectrophotometer...your cell pellet but will generally be between 250–1000 µL . The ideal lysis buffer will vary depending ...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment...January 24, 2022 Workflow Timeline Day 1: Prepare lysates, run SDS-PAGE, transfer, block, incubate with primary...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 20–200 µL multichannel...Microcentrifuge tubes, Neptune 3745.X Pipette tips, 1000 µL, VWR 76322-154 Pipette tips, 10 µL, VWR 76322...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment...instead of room temperature or 37 °C. The protocol notes when there are different incubation options available...empirically determined. If possible, run an initial test with serially diluted primary antibody and determine...Equipment Spectrophotometer compatible with 96-well plates 1–10 µL single channel pipette 2–20 µL single channel...VWR 76322-150 Pipette tips, 20 µL, VWR 76322-134 Pipettes, 10 mL, VWR 89130-898 1 L polystyrene bottle, ...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 1–10 µL multichannel pipette...calculate the titer for each sample: $$T = {R*C*D*1000\over V}$$ Where: T = Titer, GC/mL R = Reaction volume...primers, and probe on ice before use. Wipe down all pipettes and surfaces with 10% bleach. Safety Warnings ...PCR tubes, Axygen, PCR-02-FCP-C ddPCR 96-well PCR plates, Bio-Rad, 12001925 48-well dilution plate, Bio-... 1X PCR buffer (1:25,600,000) Use multichannel pipettes for the dilution series. For dilutions 1–5, use...reduce NTC values, we recommend wiping down all pipettes and equipment with 10% bleach prior to use and...

...when using highly competent cells. If you used 100-1000 ng of total DNA in a ligation you will often get... on ice (approximately 20-30 mins). Remove agar plates (containing the appropriate antibiotic ) from storage...that there isn't too much liquid media on the agar plates. If the agar plate doesn't dry adequately before...the liquid and won't grow in colonies. Incubate plates at 37°C overnight. Tips and FAQ How can I save ...access to an electroporator and the appropriate cuvettes. Follow the manufacturer's instructions for each...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...lots of FBS can promote or inhibit transfection. Test a variety of brands and lots of FBS to find one ...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Reagents 1X PBS Microcentrifuge...of our protocols supports reproducibility and accelerates science. Here, we list the specific equipment... Equipment Pipette controller Pipette tips and pipettes Rocking platform Tweezers Fluorescent microscope...

...Autoclavable flasks Sterile pipettes Ice bucket to hold antibiotic Antibiotic at 1000 x concentration disolved...number of plates you’d like to pour. For example: Because we’d like to make 20 plates, and our plates can hold... and negative test results. Sample Data In all cases below (-) indicates that the tested strain is not... Protocols Pouring LB Agar Plates Pouring LB Agar Plates You may...Sterile H 2 O Sterile plates of your desired size - we usually use 60 mm x 15 mm plates which can hold 5-10...colonies, we recommend using larger plates. Many labs use 100 mm x 15 mm plates which can hold 10 - 15 mL of ...appropriate number of plates and stack them on your lab bench. Label the plates with the date and the ...

...middle is hundreds, and the third is tens. Therefore, 1000 µL would read as 100 from top to bottom (as shown... single channel pipettes. These come in standard sizes: P2, P10, P20, P200, and P1000. The table shows...display on these pipettes will also have small tick marks at the bottom. For the P1000, this represents...video for tips on pipetting in the lab. Equipment Pipettes Pipette tips Waste container Containers to hold...Anatomy of a Pipette This front view photo of a P1000 pipette shows the plunger at the top, which is used...tip off of the pipette. This side view photo of a P1000 pipette shows the volume adjustment ring right below...Tips Although there are many different types of pipettes that have different mechanisms to disperse liquids...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...lots of FBS can promote or inhibit transfection. Test a variety of brands and lots of FBS to find one ...solution through a 0.22 μm filter to remove any precipitates. Stain each well with 1 mL of 0.1% crystal violet...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...transiently-transfected cells, increases reproducibility, as it eliminates the variation associated with repeated transient...and FBS lots can promote or inhibit transfection. Test a variety of brands and FBS lot numbers to find ... expanded, prepare cell stocks and/or harvest to test for protein expression. Typically, cells transduced...levels of the transgene may have reduced growth rates, especially if that transgene is toxic. Eventually...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 1–10 µL multichannel pipette...dilution is appropriate). Quality of duplicates: Exclude duplicates from analyses if there is more than...-log of the expected titer of the samples being tested. Always include a No Template Control (NTC), i....small aliquot of each standard (enough for 1 or 2 plates) and store at -20 °C. Once a standard is thawed...the assay multiple plasmids containing ITR were tested. Plasmid #59462 is one plasmid that gave reliable...consistent results. Use the recommended plasmid, or test multiple plasmids to find a suitable one. Some labs...95 uL 20X 200X Dilution 3 20 uL Dil. 2 80 uL 5X 1000X Dilution 4 20 uL Dil. 3 80 uL 5X 5000X Dilution ...

...MaxiPreps (Qiagen HiSpeed Max, Catalog #12663) Tips (1000 µL, 200 µL, 10 µL) Bacti Cell Spreaders (VWR, 60828... bioassay plates and remove bacteria. Use one scraper for all plates. Use two 10 mL pipettes (one for ...Antibiotic 245 mm bioassay plates (Molecular Devices, X602) Pro-Tip Pour these plates at least one day in advance...Agar + Antibiotic plates. Prewarm 12 mL recovery media at 37 °C (for at least 15 minutes). Prewarm 3X LB... Agar + Antibiotic plates at 37 °C. Prewarm 8X LB Agar + Antibiotic Bioassay plates. Prechill Micropulser...Micropulser cuvettes on ice. Thaw 4 tubes of electrocompetent cells on ice for 15-20 minutes or until completely...transformed cells on each of the eight bioassay plates (two plates per tube). Distribute evenly with a sterile...

...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 2–50 µL multichannel pipette...mixes and samples on ice before use. Wipe down all pipettes and surfaces with 10% bleach. Safety Warnings ...PCR tubes, Axygen, PCR-02-FCP-C ddPCR 96-well PCR plates, Bio-Rad, 12001925 Pierceable foil heat seal, Bio-Rad...Cell Harvest Gently aspirate media from the 6-well plates. Be sure to use a new aspirating pipette for each...and Troubleshooting We recommend wiping down all pipettes and equipment with 10% bleach prior to use and...