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Addgene

Enter a sequence to perform a BLAST-based search of the Addgene plasmid sequence database.

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We narrowed to 23 results for: SUP

Showing: 1 - 20 of 23 results

  1. Handling Plasmids from Addgene - Purifying Plasmid DNA

    Type
    Protocol
    ...precipitated DNA. Supernatant contains residues, salts, and water. Pour out the supernatant in the sink. Open...aliquot it into several tubes/bottles. Remove the supernatant and resuspend the bacteria in buffer. Note: This...centrifugation, and remove the plasmid-containing supernatant. Add either ethanol or isopropanol to precipitate...microfuge tube at 10,000 g for 30 sec. Pour off the supernatant, being careful not to disturb the bacterial pellet...salt and other extra particles from solutions. Supernatant contains the plasmid DNA separated from bacterial...bacterial chromosomes. Collect the supernatant into a new tube by pipetting or carefully pouring. Optional...Optional: Add 5 μL of 2 mg/mL RNase A to the supernatant in the new tube and incubate at 37 °C for 5 min. Note...

  2. Transfection for Recombinant Antibodies

    Type
    Protocol
    ...and valproic acid supplementation During the 24–144 h post-transfection, supplement the flask with 4% ...protease inhibitor cocktail to the supernatant. The supernatant containing the recombinant antibody can be used...from the supernatant (see our Recombinant Antibody Purification Protocol ). Store the supernatant at 4 °...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. General Considerations...culture volume. At 72-96 h post-transfection, supplement the flask with 3.75 mM valproic acid. Example... at 3100 x g to pellet the cells. Filter the supernatant through a 0.45 µm PES filter. Add 1X protease...

  3. Affinity Purification of Recombinant Antibodies with Protein A or Protein G

    Type
    Protocol
    ...purify recombinant antibodies from cell culture supernatant using Protein A or Protein G columns....purify recombinant antibodies from cell culture supernatant using Protein A or Protein G columns. Sharing...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. Safety Warnings Sodium...chromatography Equilibrate filtered tissue culture supernatant containing the recombinant antibody to room temperature...done. We typically start with about 250 mL of supernatant and add 250 µL of 100X protease inhibitor cocktail... A/G binding buffer to 1 part tissue culture supernatant and mix well. Uncap the Protein G Gravitrap or...

  4. Lentivirus ddPCR Titration

    Type
    Protocol
    ...Designate the sample name, experiment type, supermix type (ddPCR Supermix for Probes), the target names and target...Biology Grade Water, Hyclone, SH30538.02 ddPCR Supermix for Probes no dUTP, Bio-Rad,1863023 Droplet generation...Centrifuge for 5 min at 100 x g . Gently aspirate supernatant. Wash cell pellets in 500 µL PBS. Centrifuge ...Centrifuge for 5 min at 100 x g . Gently aspirate supernatant. Extract genomic DNA according to the GeneJet Genomic...Safety Cabinet (BSC). Make sure that the BSC is supplied with sufficient pipette tips. Pre-warm the 96-...Volume 9X Volume Final Concentrations 2X ddPCR Supermix for Probes, no dUTP 10 µL 90 µL 1X 20X RRE target...place the PCR plate with the foil onto the metal support block. Place the block in the plate sealer and ...

  5. Western Blot

    Type
    Protocol
    ...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. General Guidelines ...Spectrophotometer Heat block Mini gel tank chamber Power supply iBlot 2 Gel Transfer Device Roller Spatula Platform...6 cells for 5 min at 100 x g . Gently remove supernatant. Resuspend cell pellet in 1 mL of 1X PBS and ...Centrifuge for 5 min at 100 x g . Carefully remove supernatant. Resuspend cell pellet in appropriate volume ... for 15 min at 14,000 x g at 4 °C . Transfer supernatant to a clean microcentrifuge tube. Use the lysate...tank and plug the electrode cords into the power supply. Run the gel at 100 V for 10–15 min or until the...

  6. Coomassie Purity Stain of Recombinant Antibodies

    Type
    Protocol
    ...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. Last Update: June 14...controller Microcentrifuge Electrophoresis chamber Power supply Rocking platform Fume hood Metal spatula Razor ...the electrophoresis chamber and attach to a power supply. Run the gel at 150 V for 1 h . Pro-Tip If the ...“smile”, reduce the voltage. Turn off the power supply and unplug the electrophoresis chamber. Remove ... but some labs use unpurified tissue culture supernatant routinely without any issues. Plot the mg/mL ...

  7. AAV ddPCR Titration

    Type
    Protocol
    ...Designate the sample name, experiment type, supermix type (ddPCR Supermix for Probes), the target names and target...PCR Buffer, Applied Biosystems, N8080129 ddPCR Supermix for Probes no dUTP, Bio-Rad,1863023 10% Pluronic... the BSC to be used for droplet generation are supplied with sufficient pipette tips and reagent reservoirs... Volume 9X Volume Final Concentration 2X ddPCR Supermix for Probes, no dUTP 10 µL 90 µL 1X 10 µM ITR probe...place the PCR plate with the foil onto the metal support block. Place the block in the plate sealer and ...

  8. AAV Production in HEK293 Cells

    Type
    Protocol
    ...on ice and transfer supernatant to a sterile 500 mL bottle. Process the supernatant as follows: Filter ... Add 25 mL of PEG solution to each 100 mL of supernatant. Split into 2 x 500 mL sterile bottles as needed... at 3900 rpm for 15 min at 4 °C. Discard the supernatant and resuspend the pellets in a total of 5 mL ...now ready to purify your prep . The clarified supernatant can be kept overnight at 4 °C before proceeding...

  9. Kit Free RNA Extraction

    Type
    Protocol
    ...minutes at 10,000 x g at 4°C and discard the supernatant. There should be a gel-like white pellet of total...minutes at 10,000 x g at 4 °C and remove the supernatant. Remove as much of the ethanol wash as possible...

  10. Lentivirus Production

    Type
    Protocol
    ...harvesting. Filter supernatant through a 0.45 μm PES filter. The viral supernatant can be stored at 4 ...at 4 °C between harvest. Centrifuge the viral supernatant at 2100 rcf for 5 minutes to pellet any packaging...

  11. Centrifugation

    Type
    Protocol
    ...sample while the liquid portion is known as the supernatant. Depending on your experiment, you may be required...required to save the pellet, the supernatant, or both. Conclusion Centrifuges are important tools for many...

  12. Immunocytochemistry

    Type
    Protocol
    ...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. Last Update: January...

  14. Pouring LB Agar Plates

    Type
    Protocol
    ... not supposed to be resistant to the antibiotic, (+) indicates that the tested strain is supposed to be...

  15. Antibody Validation Using the Indirect ELISA Method

    Type
    Protocol
    ...that sharing the full details of our protocols supports reproducibility and accelerates science. Here,...information is solely for transparency intended to support reproducibility in science. General Considerations...

  17. Fluorescence Titering Assay

    Type
    Protocol
    ... It is not recommended to subject lentiviral supernatants to multiple freeze-thaw cycles. Procedure Seed...

  19. Colony Formation Titering Assay

    Type
    Protocol
    ...lines. It is not recommended that lentiviral supernatants be subjected to multiple freeze-thaw cycles....
Showing: 1 - 20 of 23 results