3.1 MicrowaveAssisted Drying
of Gametes
A schematic of the experimental setup used for drying gametes is
shown in Fig. 5. The microwave-assisted drying processing and the
determination of end moisture contents of samples need to be
performed in a dry chamber with a controlled RH of ~11%. The
fan of the microwave can be disabled in order to reduce the uncontrolled effects of convective drying. The threshold temperature
limit of the instrument should be set high (100
C) to avoid
power interruptions during characterization studies. In order to
observe individual sample temperatures, a handheld IR temperature sensor can be used to immediately measure temperatures after
drying (note: the microwave oven door must first be opened; thus,
associated delays could result in a difference between real and
measured temperature). When filter paper is used as a substrate
for samples, irreversible temperature-sensitive labels, placed
between filter paper and sample holder, can be used to monitor
maximum sample temperature during drying.
3.1.1 Assessment
of Microwave-Assisted
Desorption Kinetics
of Trehalose Buffer
Solution
1. In order to prescribe specific drying end points for storage
studies, a reference drying curve needs to be established for a
40 μL droplet of trehalose in Tris-EDTA buffer solutions
(1.1 M used for oocytes, 0.3 M used for sperm) using a customized microwave (CEM SAM 255, Matthews, NC) operating
at 20% microwave power (see Note 3).
2. The end moisture content should be determined at 5 min
intervals of dehydration processing for up to 50 min, in a
controlled RH of 11.0 Æ 0.6%.
3. Karl Fisher titration (V20S, Mettler-Toledo, Columbus, OH)
can be used to determine the water content of samples,
expressed as grams of H 2 O per gram of dry weight
(g H 2 O/g DW) (see Note 4).
Table 1
Special materials needed for drying of oocytes and sperm
Oocytes
Sperm
Substrate
Filter papers
(d ¼ 13 mm)
Glass coverslips
(d ¼ 13 mm)
Storage
5 mL glass vials
–
Culture medium
Minimum essential medium
HEPES solution
L-Glutamine
Sodium pyruvate
Penicillin and streptomycin
Bovine serum albumin
Hams-HEPES media
Drying Technology for Preservation of Biologics
213
of Gametes
A schematic of the experimental setup used for drying gametes is
shown in Fig. 5. The microwave-assisted drying processing and the
determination of end moisture contents of samples need to be
performed in a dry chamber with a controlled RH of ~11%. The
fan of the microwave can be disabled in order to reduce the uncontrolled effects of convective drying. The threshold temperature
limit of the instrument should be set high (100
C) to avoid
power interruptions during characterization studies. In order to
observe individual sample temperatures, a handheld IR temperature sensor can be used to immediately measure temperatures after
drying (note: the microwave oven door must first be opened; thus,
associated delays could result in a difference between real and
measured temperature). When filter paper is used as a substrate
for samples, irreversible temperature-sensitive labels, placed
between filter paper and sample holder, can be used to monitor
maximum sample temperature during drying.
3.1.1 Assessment
of Microwave-Assisted
Desorption Kinetics
of Trehalose Buffer
Solution
1. In order to prescribe specific drying end points for storage
studies, a reference drying curve needs to be established for a
40 μL droplet of trehalose in Tris-EDTA buffer solutions
(1.1 M used for oocytes, 0.3 M used for sperm) using a customized microwave (CEM SAM 255, Matthews, NC) operating
at 20% microwave power (see Note 3).
2. The end moisture content should be determined at 5 min
intervals of dehydration processing for up to 50 min, in a
controlled RH of 11.0 Æ 0.6%.
3. Karl Fisher titration (V20S, Mettler-Toledo, Columbus, OH)
can be used to determine the water content of samples,
expressed as grams of H 2 O per gram of dry weight
(g H 2 O/g DW) (see Note 4).
Table 1
Special materials needed for drying of oocytes and sperm
Oocytes
Sperm
Substrate
Filter papers
(d ¼ 13 mm)
Glass coverslips
(d ¼ 13 mm)
Storage
5 mL glass vials
–
Culture medium
Minimum essential medium
HEPES solution
L-Glutamine
Sodium pyruvate
Penicillin and streptomycin
Bovine serum albumin
Hams-HEPES media
Drying Technology for Preservation of Biologics
213