A Technique for Magnetic Glass Particle-Mediated Nucleic Acid Extraction from Blood
Transcription
Take a tube containing freshly drawn human blood and add a lysis buffer.
The detergent in the buffer permeabilizes the cell membrane, lysing the cells and releasing intracellular nucleic acids, or NAs, and nucleases.
Chaotropic agents in the buffer denature nucleases, preventing NA degradation.
Add magnetic glass particles or MGPs — silica-coated magnetic particles — and mix thoroughly.
The buffer's low pH protonates silica, rendering a positive charge on the MGPs' surface.
Negatively charged NAs bind electrostatically to the charged surface, forming an NA-MGP complex.
Under a magnetic field, NA-MGP complexes collect at the side of the tube.
Discard the supernatant containing cell debris and proteins.
Perform multiple washing steps by resuspending the NA-MGP complexes in a wash buffer, applying a magnetic field to collect the complexes, and discarding the supernatant with the remaining contaminants.
Resuspend complexes in an elution buffer. The high pH disrupts NA-MGP interaction, releasing the NAs. The eluted NAs are ready for downstream assays.
