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PVA-DABCO Immunofluorescence Mounting Solution

This document provides instructions for making a PVA-DABCO coverslipping solution for mounting immunofluorescence preparations. The solution uses glycerol as the mounting medium and contains an anti-fading reagent, PVA, and DABCO. The coverslips can be removed by soaking in buffer and the slides stored at 4°C or -20°C for at least 5 years. The document then provides a detailed recipe and instructions for making 25ml of a 2.5% PVA-DABCO solution.

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611 views2 pages

PVA-DABCO Immunofluorescence Mounting Solution

This document provides instructions for making a PVA-DABCO coverslipping solution for mounting immunofluorescence preparations. The solution uses glycerol as the mounting medium and contains an anti-fading reagent, PVA, and DABCO. The coverslips can be removed by soaking in buffer and the slides stored at 4°C or -20°C for at least 5 years. The document then provides a detailed recipe and instructions for making 25ml of a 2.5% PVA-DABCO solution.

Uploaded by

Dgek London
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© Attribution Non-Commercial (BY-NC)
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as PDF, TXT or read online on Scribd

Peterson lab

Dept. Neuroscience

PVA-DABCO Coverslipping Solution for Immunofluorescence


This solution is a glycerol based mounting medium containing an anti-fading reagent for use with immunofluorescence preparations. The same solution may be used with or without DABCO for aqueous mounts of material for brightfield microscopy. The advantage of this solution is that the medium at the perimeter of the coverslip polymerizes upon contact with air forming a strong seal. There is no need to seal with nail polish. The coverslips can be removed by soaking the slide in buffer. Mount stained sections on glass slides and let briefly air dry (less than one minute) or if using Lab-Tek chamber slides, remove the chamber and tip to remove excess buffer. Apply 75 l of PVA-DABCO solution for use with a 50 mm coverslip. Use caution with pipetting as air bubbles are easily introduced if the solution is aspirated in any way. I suggest using a repeating pipetteman. Lay the coverslipped slides flat overnight until polymerization is complete. Afterwards, the coverslips may be cleaned with solvents to remove immersion oil and slides can be stored upright in normal slide storage boxes. Store slides in slide boxes at 4C or -20C and preparations should remain usable for at least five years, the longest time point evaluated to date. Daniel A. Peterson, Ph.D. March 2001 For 25 ml of 2.5% PVA-DABCO Make 25 ml of PVA-DABCO in 50 ml culture tube to allow ample room for mixing. 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. Using a 50 ml culture tube, weigh 6 gm of glycerol into the culture tube on the balance Add 2.4 gm of PVA Mix well by repeatedly inverting the tube until the PVA is coated with glycerol Add 6 ml of distilled water and repeat mixing until relatively uniform Mix overnight using a rotator at room temperature Add 12 ml of 0.2 M Tris-HCl at pH8-8.5 Heat to 50C in a water bath with mixing (approximately 30 min) Add 0.625 gm of DABCO and mix well Centrifuge at 5000g for 15 min Remove supernatant, aliquot (suggest 1 ml), and store at -20C

The solution may be kept up to 6 months at -20C and up to one week at 4C before becoming milky. Do not refreeze.

Peterson lab Stock materials PVA-polyvinyl alcohol DABCO- 1,4 diazabicyclo [2.2.2]octane Tris-HCl Glycerol Vendor Sigma (P8136) Sigma (D2522) Sigma (T3253) Fisher G33-1

Dept. Neuroscience Location Chemical Shelf Chemical Fridge Chemical Shelf Chemical Shelf

To make 0.2N Tris HCl: 500 ml ddH2O 15.76g Tris HCl add7 pellets NaOH adjust pH to 8.0-8.5

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