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Rapid Miniprep Protocol For Eppendorf

  1. Spin cells to pellet down and discard supernant (use 1.5 ml of cells).
  2. Add 100ul Tris/EDTA/Glucose (TEG) and resuspend cells.
  3. Add 200ul SDS/NaOH immediately and tap tubes.
  4. Add 150ul K/Na acetate (1-3 min after lysis step).
  5. Shake tubes vigorously by inverting several times.
  6. Spin tubes 14000g at RT/5 min.
  7. Transfer the supernatant to a fresh tube.
  8. Add = vol (~450ul) phenol/chloroform, and vortex. (Or use Progenius method, listed below (in part).)
  9. Spin tubes 14000g RT/3 min.
  10. Transfer supernatant to fresh tubes and add 2 vol 100% ethanol (~700ul), invert tube several times then spin 14000g RT/20 min.
  11. Remove supernatant and add 70% ethanol, vortex then spin RT/5 min.
  12. Remove supernatant, dry pellet (speedvac med heat setting/5 min).
  13. Resuspend pellet in 45ml T10E 1 5ml Rnase (20ug/ml).
  14. Heat tube at 55 0 /5 min.
  15. Cut appropraite amount or store at -20 0 C.
  • TEG(50ml) 5 ml 0.5M Glucose, 1 ml 0.5M EDTA, 2.5 ml 0.5M Tris pH 8.0, 41.5 ml H 2 O
  • K/Na Acetate (50ml) 30ml 5M K Acetate, 5.75ml glacial acetic acid, 14.25ml H 2 O

Progenius Method (in part):

  1. Add 500ml NaI sol n ,100ml TE, 5ul silica to the supernatant of each tube (Silica stock needs to be vortex for 5-10min).
  2. Shake gently for 5-10 min.
  3. Centrifuge for 30 sec, Aspirate supernatant and discard.
  4. Preform 1ml ethanol wash (Progenius Kit). Do twice.
  5. To elute:
    • use 25ul H 2 O
    • heat at 55C for 2-5 min then centrifuge for 30 sec.
    • Remove supernatant to fresh eppendorf.
    • Relute pellet in 25ul H 2 O, heat at 55C for 2-5 min then centrifuge for 30 sec.
    • Again remove supernatant and place with previously removed supernatant. (total 50ul).