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...Pipette controller Benchtop centrifuge compatible with 50 mL conical tubes NanoDrop spectrophotometer 37 °...Immunocytochemistry Protocol Western Blot Protocol Introduction This protocol describes how to affinity purify...concentration of each fraction on the NanoDrop Spectrophotometer using the A280 IgG setting. Combine all eluates...concentration of the pooled sample on the NanoDrop Spectrophotometer using the A280 IgG setting. If the concentration...Determine antibody concentration on the NanoDrop Spectrophotometer. Dilute antibody to 1 mg/mL with PBS if needed...to check the concentration on the NanoDrop Spectrophotometer to see if the sample has reached the desired...Periodically check the concentration on the NanoDrop Spectrophotometer to see if the sample has reached the desired...

...of each well at 450 nm on the spectrophotometer. Use the spectrophotometer to generate a standard curve...Antibody Purification Protocol Antibody Guide Introduction This protocol describes how to run an indirect...antibody incubation and plate read Tips and Troubleshooting The optimal concentration of primary antibody...an automated ELISA plate washer. Equipment Spectrophotometer compatible with 96-well plates 1–10 µL single...Multichannel pipette reagent reservoirs Pipette controller Microcentrifuge Microplate shaker ELISA plate... antigen Purified recombinant antibody Isotype control antibody HRP-conjugated isotype-specific secondary...mix. Dilute the primary antibody and an isotype control antibody to the desired concentrations in the antibody...

...questions. Intro to the Lab Bench Basic Molecular Biology Plasmid Cloning Virus Antibodies Intro to the Lab... Lab Bench Introductory techniques designed to help you get started in the lab. Name Description (Link...Quantification Measure DNA concentration with a spectrophotometer Restriction Digests Cut and analyze plasmid...plasmid DNA with restriction enzymes Agarose Gel Electrophoresis Separate DNA by size on an agarose gel Watch...DNA using DNA ligase Bacterial Transformation Introduce DNA into a bacterial strain Watch the Video! CRISPR...Description Link to Video General Transfection Introduce plasmid DNA to mammalian cells Lentivirus Production.... Name Description Link to Video Transfection Introduce plasmid DNA to mammalian cells to produce antibodies...

...DNA Ligation Introduction Transformation is the process by which foreign DNA is introduced into a cell....should also add a positive control (many companies include a positive control plasmid with their competent...ligation of inserts to vectors than for an intact control plasmid. Incubate the competent cell/DNA mixture...transform large plasmids, it is a good idea to use electro-competent cells. Instead of relying on the heat-shock...heat-shock to cause the cells to take up the DNA, an electro-magnetic field is applied to the cell/DNA mixture... do this you will need to have access to an electroporator and the appropriate cuvettes. Follow the manufacturer's...

...and FAQ Do controls When doing ligations you should ALWAYS do a vector alone + ligase control. This will...ligation. Additional controls are encouraged, but may only be required for troubleshooting failed ligations...The following table indicates the various controls: Control Ligase Interpretation Uncut vector - Checks...know has only been thawed once before. Always do controls. See Tips and FAQ below for details. Try different...treated, that the phosphatase treatment worked. This control should, in principle, be free of colonies, but ...

...purity. This can be done with a spectrophotometer. A spectrophotometer uses the absorbance/transmission... instructions for the spectrophotometer specific to your lab. Spectrophotometer Tips Before measuring ...labs have a NanoDrop, which is a very small spectrophotometer that can accurately read DNA concentration...measuring any samples, be sure to ‘blank’ the spectrophotometer using the solution the DNA is resuspended in, ...NanoDrop. It is a good idea to re-zero any spectrophotometer between samples if they are expected to vary...

...an uncut sample of each vector to help with troubleshooting if your digests don’t look as you expected....product is the anticipated size and that your band is strong (indicating that the PCR reaction worked and that...insert. It is also important to set up negative controls in parallel. For instance, a ligation of the recipient...much less total DNA (<1ng) or if you are having trouble getting colonies, you might want to use higher ...to be very large (>10kb) you might want to use electro-competent cells instead of the more common chemically-competent...plasmid alone plate. The recipient plasmid alone control will tell you your “background” level or more specifically...see any colonies, you should conduct a positive control to ensure that your transformation worked. You ...

...Restriction Digest of Plasmid DNA Agarose Gel Electrophoresis Introduction Restriction enzymes are naturally occurring...and analyze the resulting fragments by gel electrophoresis . The pattern of the fragments on the gel can...to analyze a restriction digest. Equipment Electrophoresis chamber Pipetman Pipet tips Reagents Liquid...manufacturer's instructions) Gel loading dye Electrophoresis buffer Verifying Total Plasmid Size -OR- Insert...avoided. If you do have to do so, there is no way to control which orientation the insert is ligated into the... tricks see the Restriction Digest and Gel Electrophoresis pages....

... uncut sample of each plasmid to help with troubleshooting if your digests don’t look as you expected....insert. It is also important to set up negative controls in parallel. For instance, a ligation of the recipient...much less total DNA (<1ng) or if you are having trouble getting colonies, you might want to use higher ...to be very large (>10kb) you might want to use electro-competent cells instead of the more common chemically-competent...plasmid alone plate. The recipient plasmid alone control will tell you your “background” level or more specifically...see any colonies, you should conduct a positive control to ensure that your transformation worked. You ...depending on the number of background colonies on your control plate (the more background, the more colonies you...

...channel pipette Pipette controller Pipette tips and pipettes Spectrophotometer Heat block Mini gel tank...Protocol Recombinant Antibody Purification Protocol Introduction Western blot is a technique used to separate...ladder. Place the lid on the tank and plug the electrode cords into the power supply. Run the gel at 100... a dark room to detect the bands. Tips and Troubleshooting The optimal lysis buffer will vary depending...as expected and is specific, include a positive control sample that you know expresses the protein, such...transfected with the protein of interest, and a negative control sample that does not express the protein of interest...

...Viral Service Lentivirus Guide Virus Blog Posts Introduction This protocol describes how to use droplet digital...ribonuclease P/MRP 30 kDa subunit (RPP30) are used as a control for normalization. The dilution series outlined...wells containing the virus and the untransduced control. Pro-Tip For even seeding, prepare a batch for ...Determine the concentration of each sample on a spectrophotometer. Prepare 25 ng/µL stocks of each sample. Samples...calculate the final infectious titer. Tips and Troubleshooting We recommend wiping down all pipettes and equipment...concentration of RRE positive droplets in the untransduced control should be close to zero (A01). In this protocol...

...Protocol Recombinant Antibody Purification Protocol Introduction Immunocytochemistry is a technique that uses...Day 3-4: Fix and label cells Equipment Pipette controller Pipette tips and pipettes Rocking platform Tweezers... appropriate fluorescent filters. Tips and Troubleshooting The optimal fixation method will vary depending...functioning as expected and specific, include a positive control sample that you know expresses the protein, such...express the protein of interest, and a negative control sample such as cells that do not express the protein...

...Virus Blog Posts Mol Bio Protocols Viral Service Introduction This protocol can be used to titer lentivirus...titer of your lentiviral preparation allows you to control the multiplicity of infection (MOI) in downstream...channel pipette Ice bucket CO 2 incubator Pipet controller Hazardous waste container Reagents DMEM high ...All of the cells in the untransduced (negative) control well should be killed and no colonies should be...appropriate antibiotic. When the untransduced (negative control) cells have died and colonies are visible in the...

...Gel Purification Molecular Biology Reference Introduction This protocol describes how to extract RNA from...unlike using a p1000 tip it will give you more control of where you’re aiming your tip in the tube. Precipitate...the quality of your RNA sample(s) using a spectrophotometer (such as a Nanodrop), agarose gel, or bioanalyzer...

...gel electrophoresis, which separates DNA by their length in base pairs. Following electrophoresis, you...Restriction Digest of Plasmid DNA Agarose Gel Electrophoresis DNA Ligation Background Information Gel purification...Protocol: Gel Purification Follow the Agarose Gel Electrophoresis Protocol with the following amendments: Notes...

... Technology Protocol Agarose Gel Electrophoresis Gel electrophoresis is the standard laboratory procedure...visualization and purification. Agarose Gel Electrophoresis Protocol Purifying DNA from an Agarose Gel ...DNA fragments based on size. Following gel electrophoresis, you can cut DNA bands out of the agarose gel...

...Virus Blog Posts Mol Bio Protocols Viral Service Introduction This protocol can be used to titer lentiviral... titer of your lentiviral vector allows you to control the multiplicity of infection (MOI) in downstream...channel pipette Ice bucket CO 2 incubator Pipet controller Hazardous waste container Inverted fluorescence...dilution with one well remaining as the untransduced control). Count the cells in the remaining well, a cell...

...Purifying DNA from an Agarose Gel DNA Ligation Introduction Restriction enzyme digestion takes advantage...your restriction digest results. Equipment Electrophoresis chamber Pipetman Reagents Liquid DNA aliquot...manufacturer's instructions) Gel loading dye Electrophoresis buffer Pipet tips Procedure Select restriction...visualize the results of your digest, conduct gel electrophoresis . Tips and FAQ Restriction enzymes MUST be ...

...Using a Light Microscope Using a Light Microscope Intro to the Lab Bench Check out more protocols and videos...videos to help you get started in the lab! Introduction Microscopes are emblematic of biological research... cheap and accessible) to massive transmission electron microscopes that allow us to see cellular components...resolution when your image is in focus, which you can control using the focus knobs on the side of the microscope...

...Virus Blog Posts Mol Bio Protocols Viral Service Introduction This protocol can be used to generate stable...channel pipette Ice bucket CO 2 incubator Pipet controller Hazardous waste container Reagents DMEM high ...plate will hold 5 dilutions plus one 'no virus' control well). Perform a "reverse transduction" by seeding...selection should last at least as long as it takes the control (untransduced) cells to completely die. After that...