I am trying to pull glass pipettes for Intracranial AAV microinjection into dentate gyrus (my understanding is that they are best at 20-50um tip diameter) but I am not sure what temp./pull/velocity/time to use.
I use a Sutter Instrument Model P-1000 puller and WPI Borosilicate Glass Pipettes 1B100F-4 ( 1.0 OD, 0.58 ID)
Please let me know if anybody can share a protocol? Thanks!
Please find enclosed Sutter Technical Tip instructions in PDF format...given to me several years ago from a member of the Sutter technical team UK when I was working on Xenopus
I am using
- Narishige PE-22 puller
- Glass capillary (OD: 1mm; ID:0.78mm, Warner Instruments, G100T-4, # 640778)
I found heater, sub magnet and main magnet value of puller to 88.7℃; 20.1 and 74.8 respectively best
Higher heat (89.7℃) will result in longer (7~8mm) and finer tip (50µm) and lesser heat (88.7) will result in smaller (4~5) and larger diameter (70µm) tip . Similarly, powerful electromagnetic force, permits the production of a long and thin micro-pipette with a sharp-pointed tip.
I mostly used (70µm) tip for micro-injection and inject .200µl/5min by automatic injector (Narishige IMS-20)
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