Primary human cortical astrocytes (ScienCell) were obtained commercially and expanded in astrocyte media (ScienCell). Cells were cultured at 37°C in 5% CO2 and split at approximately 80% confluency until passage 3. Upon reaching passage 3, cells were frozen using Recovery Cell Freezing Media (Gibco). Cells were thawed prior to experiments and transfected during passage 4. Vectors encoding either intron 2 retained PSMB8 (i2R-PSMB8) or the canonical full-length transcript (FL-PSMB8) in pcDNA3.1 were obtained from GenScript, outgrown in DH5α E. coli (New England Biolabs), and midi-prepped using a Plasmid Plus Midi kit (Qiagen) per manufacturer’s instructions. Vector maps are provided as Supplementary Figures S1, S2 . Vectors were transfected using a Lonza Amaxa Nucleofector 4 device, with kit P3 and protocol DR114, at a ratio of 6 μg DNA per 106 cells. This resulted in a 50–55% transfection efficiency (Supplementary Figure S3 ).
Amaxa nucleofector 4 device
Manufactured by Lonza
The Amaxa Nucleofector 4 device is a laboratory instrument designed for the electroporation-based transfection of cells. It is used to introduce nucleic acids, such as plasmids, siRNA, and other molecules, into a variety of cell types. The device provides a controlled and efficient method for delivering these materials into the cell nucleus.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using amaxa nucleofector 4 device
1
Transfection of Astrocytes with PSMB8 Variants
2
Transfection of Primary Human Astrocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary human cortical astrocytes (ScienCell) were obtained commercially and expanded in astrocyte media (ScienCell). Cells were cultured at 37°C in 5% CO2 and split at approximately 80% confluency until passage 3. Upon reaching passage 3, cells were frozen using Recovery Cell Freezing Media (Gibco). Cells were thawed prior to experiments and transfected during passage 4. Vectors encoding either intron 2 retained PSMB8 (i2R-PSMB8) or the canonical full-length transcript (FL-PSMB8) in pcDNA3.1 were obtained from GenScript, outgrown in DH5α E. coli (New England Biolabs), and midi-prepped using a Plasmid Plus Midi kit (Qiagen) per manufacturer’s instructions. Vector maps are provided as Supplemental Figures 1 –2 . Vectors were transfected using a Lonza Amaxa Nucleofector 4 device, with kit P3 and protocol DR114, at a ratio of 6 μg DNA per 106 cells. This resulted in a 50–55% transfection efficiency (Supplemental Figure 3 ).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!