Zeiss Lightsheet Z.1, Руководство по эксплуатации

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CHAPTER 2 - SAMPLE PREPARATION
Carl Zeiss Specific Examples of Sample Preparation Lightsheet Z.1
36 000000-1790-528 02/2013
Method:
−
Choose an adult Anopheles gambiae and immobilize it by cold treatment. Melt 1.5 % LMP
agarose, aliquot 0.5 ml into a 1.5 ml Eppendorf tube. Invert the tube to mix and allow agarose to
cool to 40º C.
−
To avoid bubble formation on the insect surface that will affect imaging, the insect must be treated
either with ethanol 70 % (animal death) or using 50 % glycerol or 1 M sucrose to cover the
hydrophobic chitin surface prior to embedding (Fig. 19/
B
).
−
The syringe is prepared as previously described and filled with molten low melting point agarose
(40 °C).
−
The insect can then be inserted into the agarose cylinder and aligned using a needle or forceps
(Fig. 19/
C
).
−
The insect can then be imaged (Fig. 19/
D
).
This technique can be applied to any insect or similar type of organism possessing an exoskeleton.
Depending on the animal part to be observed the insect can be aligned differently or dissected
prior to embedding (head, wings, guts, salivary glands…).
Fig. 19 Mounting a complete adult Anopheles gambiae .
(A) The insect is paralyzed. (B) The chitin surface is treated using 70 % ethanol. (C) The insect is
positioned within the melted agarose. (D) The mounted insect is ready for imaging.