Love:Preparing poly-lysine slides: Difference between revisions
(New page: posted by Qing Han, 4.24.08 Based on several protocols available online (DeRisi Lab at UCSF, U of Wisconsin gene expression center, Hu Lab at U of Minnesota) and modified by Love Lab. ''...) |
(No difference)
|
Revision as of 08:13, 24 April 2008
posted by Qing Han, 4.24.08
Based on several protocols available online (DeRisi Lab at UCSF, U of Wisconsin gene expression center, Hu Lab at U of Minnesota) and modified by Love Lab.
Basic note:
1.Do not use powdered glove at any time.
2.Avoid dust: try to keep the glass in solution at all times, cover the mTUB when stiring, clean all the surrounding places.
3.Wear eye protection, mask, and lab coat, and be cautious when manipulating NaOH.
4.The minimal volume for covering all the glass slides in mTUB is 500 ml.
Solution:
1.Glass clean solution (500 ml): Dissolve 50 g NaOH in 200 ml H2O, then slowly add 300 ml 95% EtOH. This solution can be reuse for next time.
2.Poly-L solution (500 ml): 407 ml H2O, 50 ml filtered PBS, 43 ml poly-lysine (Sigma, Cat.# P4832 ). This solution should be store and used in the plastic container. When filtered, it can be stored in 4 degree and reused next time. When reused, additional 1-2 ml poly-lysine can be added to the old solution each time and can be repeated of 6 times.
Protocol:
1.Loading Corning Micro Slides (2947-3*1) to the rack of mBox (20 slides/rack), and put the whole rack into mTUB (with stir bar in the bottom). Carefully pour glass clean solution onto the glass until all the glass slides are submerged. Gently stir the clean solution for 2 hours or longer (do not overnight).
2.Take out the rack with mBox handle, briefly drain, and put the whole rack into another mTUB prefilled with pure H2O (operate carefully and try to decrease the time that glass slides are exposed to the air to decrease dust adsorption. Wash several times with H2O to remove NaOH (at least 5 times, 5~10 min/ time). It is best to let the glass in the water overnight at the last wash. You can test the pH of the H2O to make sure the remove of NaOH at last.
3.Remove the rack with glass slides from H2O, briefly drain, and submerge it in poly-lysine solution, gently stir for 1 h.
4.Wash slide with H2O as described above (3 times is OK).
5.Spin glass slides to dry and put in oven at least 20 min (40~50 degree).
6.Store slides in vacuum for at least 2 weeks before use. Once coated, slides are usually stable for 4 months.
