April 5, 2016 at 10:24 am #3240
Please see the details outlined below which discusses the Poly-L-Lysine protocol. The attached document also provides some additional details and example images.
– Cationic charges electrostatically binds anionic surfaces of glass and specimen
– Poly-D-Lysine may be used if there are proteases in the sample
– Use PLL stock containing polymers in the range of 70-150 kDa
– If creating own stock, make 1% (w/v) solution with ddH2O
– Specimen may flatten and morphology may be affected depending on cell type
To make poly-L-lysine coated slides:
1. Make a .1% (v/v) working solution of poly-L-lysine (PLL) with ddH2O or PBS.
It’s best if this solution is made immediately prior to use. Keep stock in -20ºC
for up to 2 years.
2. Coat cleanroom slides thoroughly (shaker would be ideal) with PLL solution
and let air dry for about 5 minutes at room temperature. Let dry in a low-dust
environment, such as a laminar flow hood. Slides will stay good stored at
4ºC for up to a year.
3. Depending on the sample, it may also be desirable to coat coverslips with
PLL as described above.
4. Retrieve slides from 4ºC immediately prior to use. The fixative appears to be
less effective if the slides are left out at room temperature.
5. Vortex sample, drop 2.0 μL or less onto the PLL-coated slide, and add a
coverslip. Depending on the sample, you may need to wait from 30 seconds
to 10 minutes for the specimen to adhere.
6. Once all movement has ceased, sample is ready for optical and
*Note: PLL-coated slides and coverslips are commercially available, however
are usually more expensive than homemade slides. It is advantageous to
purchase these slides if possible. It is easier to contaminate the homemade
slides with dust and particulates during the drying process, and the homemade
slides typically don’t last as long as the commercial slides.
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