Biology Reference
In-Depth Information
The temperature is also an important consideration as
circulation of perfusate at room temperature will create tempera-
ture gradients across the probe that may modify immediate envi-
ronment. Hence, prewarming of perfusate to body temperature
before use is recommended (
36
).
Flow rate and sampling interval are determined by the sample size
required for analysis and the temporal resolution of molecules
being sampled. Flow rate is directly proportional to recovery; the
slower the fl ow rate greater the recovery. For cerebral microdialy-
sis, perfusion fl ow-rate typically ranges from 0.1 to 5
4.2.3. Flow Rate
and Sampling Interval
μ
l/min
(Table
1
) (
27-30
).
The data obtained from microdialysis measurements is semiquanti-
tative and is expressed as relative recovery. The term relative recov-
ery is defi ned as the ratio between substance concentrations outside
the probe to the concentration in the dialysate (
37
). Recovery is
infl uenced by the size of the membrane pore size, membrane area,
fl ow rate (a longer membrane and a lower fl ow rate will give a
higher recovery) and diffusion speed of the substance (
38
).
Recovery of each substance should be measured in vitro and men-
tioned when presenting data (see Methods).
4.2.4. Relative Recovery
A number of methods can be used for analyzing microdialysis sam-
ples. Two commonly used methods are high performance liquid
chromatography (HPLC) analysis with detector suitable for the
substances of interest (
29, 30, 35, 39
) and CMA micro analyzer
(CMA 600; Solon) (
28, 40, 41
). A good correlation is found
between the results of CMA600 analyzer and HPLC (
42
). In
recent years, easy-to-use CMA analyzer has gained much recogni-
tion for detecting markers of glucose metabolism, glutamate, and
glycerol levels in dialysate from humans after SAH (
43-45
).
4.2.5. Sample Analysis
Insertion of microdialysis probe into the brain causes injury that
may evoke infl ammation. Upon histological examination localized
tissue trauma at the site of probe insertion is observed as clotted
blood, macrophage infi ltration, and tissue retraction. In addition,
in a naive animal probe insertion trauma is also observed as greater
initial values of glucose, lactate, and glutamate in dialysate that
tapers off with time. Hence, preliminary experiments sampling
dialysate for energy-related metabolites and glutamate are recom-
mended to determine the time required for tissue to recover from
probe insertion trauma before sampling towards an experiment
could be started. A 24 h time elapse between probe implantation
and sampling is recommended (
46
). However, this time varies
depending upon the surgeon's hands and technique. We and oth-
ers have found 120 min are suffi cient to recover cerebral metabo-
lites to basal level after insertion of microdialysis probe in rats
(
28-30, 39, 47
).
4.2.6. Surgical Trauma
by Probe Implantation
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