Instruments and Materials Used in Preparation of Microscope Slides for Powdery Mildew Fungi Identification
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Step #1 - Take a clean microscope slide and add 1 drop of 5%
KOH to the center of the slide. The 5% KOH solution helps to rehydrate
the cleistothecia without rupturing them.
Step#2 - Cleistothecium Transfer Technique #1 - The use
of a dissection microscope allows you to select desired individual cleistothecia
from a collected sample. Under the dissection microscope, gently lift off
individual cleistothecia with the scalpel blade and dissection probe and
drop onto 5% KOH drop on center of slide. Hint: It may help to have the
scalpel tip wet with the 5% KOH solution.
Cleistothecial Transfer Technique #2 - If a dissecting microscope
is unavailable, a hand lens can be used to identify the location of cleistothecia
on the sample. This area can be gently scraped over the 5% KOH drop to
allow cleistothecia to fall from the sample to the slide.
Step #3 - Gently place the coverslip over the 5% KOH drop and
cleistothecium. Apply the coverslip by placing one edge down first and
then lowering the rest of the coverslip down slowly, to reduce the number
of air bubbles that will be trapped between the microscope slide and the
coverslip.
Step #4 - If air bubbles do occur, the prepared microscope slide
can be gently waved over an alcohol burner, approximately six inches from
the flame for only a few seconds. DO NOT ALLOW THE SAMPLE TO BOIL!!!
Step
#5 - Place the prepared slide under the compound microscope and use
the lowest power magnification to focus in on the sample. Locate the cleistothecia
(100x magnification is usually sufficient to do this). Refocus and increase
magnification as necessary.
Step #6 - Observe appendages of the cleistothecia and refer to a key of the Genera of Powdery Mildew.
( If identification is necessary through an asci count, remove slide from microscope and gently press coverslip down with dissection probe to rupture the cleistothecia and release the asci. Note: Be careful not to crack the coverslip.)
Step #7 - Repeat step #5. Locate ruptured cleistothecia and
observe asci numbers. Refer back to Genera key of Powdery Mildew for further
identification
Step #8 - Dispose of all glass appropriately.