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Section 1. Clinical Medicine
https://doi.org/10.29013/ELBLS-23-2-3-8
Griselda Korgari, Department of Technical Medical Laboratory and Imaging, Faculty of Medical Technical Sciences, Aldent University, Tirane, Albania
THE STABILITY OF GLUCOSE AND TRIGLYCERIDE ANALYSIS IN GEL TUBES AT 2-8 oC
Abstract
Introduction: Studies have shown that there are important, significant relationships between the values of different analytes with different physical variables such as temperature, measurement time, and centrifugation time. In this paper, the relationship that may exist between the concentration values of two analytes such as blood glucose and triglycerides with the storage temperature of the samples has been studied.
This study aimed to determine the analyte stability of venous blood samples in serum gel tubes stored at 2-8 0C.
Methods: 30 healthy adult volunteers take part in the study. Blood was collected in tubes with gel (Clot activator). All samples were allowed to clot at room temperature for 30 min. They were centri-fuged CAPP CENTRIFUGE at 3500 RPM and 2 analytes were analyzed: Triglycerides and Fasting Blood Glucose. These values were determined as control values. Serums were stored at 2-8 oC. Measurements were repeated after 1 week.
Results: 18 men and 12 women participated in the study. Triglycerides control values range from 54-149 mg/dl average (86.58mg/dL).
Fasting blood glucose values were in the range of 77-91.8 mg/dl, with an average value (of 80.23 mg/dL).
After keeping the serum at a temperature of 2-8 oC for 1 week, the following results were obtained:
• Triglyceride values ranged from 60.3-117 mg/dl, mean of value (92.25 mg/dl);
• Glucose values ranged from 53-124.4mg/dl on average (83.7 mg/dL).
Conclusions: There is a statistically significant relationship between the storage temperature of the sample and the concentration of the Triglyceride analyte (p - 0.00), while for the comparison of Glucose control and Glucose values after one week, they are not statistically significant (p = 0.2).
Keywords: Gel tube, Triglyceride, fasting blood glucose, temperature.
Introduction
Laboratory examinations have an important role in clinical diagnoses. This has made some tests part of the check-up routine. Tests such as fats and blood sugar have a role both in the diagnosis and in the prognosis of a certain pathology, so often for these tests, there is a need to repeat the measurements.
Most biochemical tests are performed on serum samples, that is, in gel tubes (Clot activator) Some studies show that these tubes maintain the stability of the values without the need to divide the samples into other tubes, but there are also studies that prove the opposite. The purpose of this study is precisely to see the stability of the values of the analyte Glycemia and Triglyceride in the tubes with gel but exposed to the temperature variable
2. Materials and methods
The study involved 30 (thirty) volunteers, who were healthy adults, not anemic, and they were not on an anticoagulant therapy. Twelve female and 18 men volunteers between the age of19 and 22 took part of this study. Five milliliters of venous blood were drown from each volunteer using a vacuum machine and gel tubes (Clot activator)
2.1 Collection and processing of samples
Each volunteer underwent a phlebotomy procedure for blood collection. Blood was collected from forearm veins using a 21G vacuum system. All samples were identified and homogenized at least five times. Each sample was allowed to be set for 30 minutes at 18-25 °C. All tubes were centrifuged at 1500g/3500 RPM for 10 minutes at 18-25 °C.
2.2 Analysis of samples
All tubes were evaluated for hemolysis, icterus, and lipemia. The CYANSMART device (ELITEK) was used to analyze the samples. Triglyceride and glucose reagents were kept at a temperature of 18-25 °C for 15-20 minutes. After each test, a calibration and control procedure was performed.
The principle of the Glycemic measurement method is: Glucose oxidase
^METHOD & PRINCIPLE <4>
Enzymatic I PAP - End Point.
Glucose +■ 0,
auww cmdur
-> Gluconic acid + HX>}
Ptnmjtit
2H.O-. + Phenol + 4-Aminoantipyrine->
Quinoneimine + 414,0
Figure 1. The principle of the glucose method
Manual procedure
• Wavelength 505 nm;
• Optical path: 1cm;
• Sample/ Reagent ratio: 1:100;
• Temperature: 37 °C Read against reagent blank.
Blank Calibration Test
Reagent R 1000 M 1000 M 1000yl
Distilet water 10 M —
Standart/ Calibrator - 10 M —
Sample - — 10 Ml
Mix and read the absorbances (A) after an incubacion of 10 minutes
METHOD 1,1
Enzymatic-colorimetric. End point, PRINCIPLE <*
Enzymatic determination of triglycerides according to the following reactions:
Upoprofeui Lipase Triglycerides + H^O->■ Glycerol + Fatly adds
GtycemJ kinase Glycerol + ATP -GlyceioW-
Phosphate+ ADP
GPO
Glycerol-3 FTiosphate + O,--0 HA- P t H^
Peroxidase
H20, + 4-AAP + j^Chioropfenoi ->
Quinoneimine
GPO = Glycerols-phosphate oxidase DHA-P = Oihydroxyacetone-P 4-AAP * Amlno-4-antipyrine
Figure 2. The principle of the Triglicerid method
The principle of the method of measuring triglycerides vs Triglyceride Oxidase. Both spectrophotometry methods are END points. Manual procedure
• Wavelength 505 nm;
• Optical path: 1cm;
• Sample/ Reagent ratio: 1:100;
• Temperature: 37 °C; Read against reagent blank.
Standart/ Calibrator - 10 ^l -
Sample - - 10 ^l
Blank Calibration Test
Reagent R 1000 ^l 1000 ^l 1000^l
Distilet water 10 ^l -
Mix and read the absorbances (A) after an incubación of 10 minutes
All samples were measured at the beginning and their values were considered control values. Afterward, the gel tubes were stored in a refrigerator at a temperature of 2-8 °C. After 1 week, these tubes were left at room temperature for 10-15 min, re-centrifuged and re-measured, maintaining the same calibration and control conditions.
Figure 3. Semi-automatic biochemistry analyzer CYANSmart
3. Results of the study statistical analysis showed that there is a statistically
All measurement values were entered into significant relationship between the concentrations the SPSS version 21 program and processed. The of triglycerides and the temperature variable. Table 1. - Indicates the statistical relationship between Tg concentrations. According to the statistical processing, we have a statistically significant relationship (P = 0000) between the concentration of Tg control and the concentration of Tg after 1 week at a temperature of 2-8 oC
Paired Samples Test
Paired Differences t df Sig. (2-tailed)
Mean Std. Deviation Std. Error Mean 95% Confie val of the lence Inter-Difference
Lower Upper
Pair 1 The concentration of Tg control Tg concentration after 1 week -5.67000 7.87594 1.43794 -8.61092 -2.72908 -3.943 29 0.000
Figurel. Shows the scatter blot of the relationship between the Tg concentration in a 1-week
time difference at a temperature of 2-8 oC. As can be seen from the graph obtained from SPSS VERSION21, there is a linear distribution of values, which shows that there is a positive statistical relationship between the concentration of triglyceride and the temperature variable
Additionally, it was demonstrated that there is y = 12.7 + 0.92 x. It was demonstrated that there is
no correlation between glucose concentrations and no correlation between glucose concentrations and
temperature in the region 2-8 °C. The linearity is temperature in the region 2-8 °C. R = 0.873 and the equation obtained from excel is
Table 2. - Shows the statistical relationship between fasting glucose concentrations. According to the statistical analysis, there is no statistically significant relationship (P=0.2) between the control glucose concentration and the glucose concentration after 1-week storage at a temperature of 2-8 oC
Paired Samples Test
Paired Differences t df Sig.
95% Confidence (2-tailed)
Mean Std. De- Std. Er- Interval of the
viation ror Mean Difference
Lower Upper
Glucose con-
Pair 1 centration after
1 week - con- 3.46933 15.71995 2.87006 -2.40059 9.33926 1.209 29 0.237
trol glucose concentration
Figure. 2 shows the scatter blot of the relationship between the end of Glycemia in a time difference of 1 week at a temperature of 2-8 oC. As can be seen from the graph obtained from SPSS VERSION21, there is a non-linear distribution of values, which shows that there is no positive statistical relationship between the concentration of glucose and the temperature variable, so the concentration changes. This means that the gel tube does not adequately maintain the glucose concentration
4. Discussions cess of cellular energy production, a process which
From the results of the work, different interpre- is stimulated by the temperature factor. the effect
tations and opinions arise which will be analyzed in the following works for the measurement of analytes. The results of the work showed the statistical relationship that existed between the temperature variable and Triglycerides, and the lack of a statistically significant relationship between the glucose concentration and the temperature variable, which makes us analyze what would happen if maintaining the same measurement procedures but extending The time the samples stay in the temperature range of 2-8 °C.
It is known that the gel tubes can maintain the stability of the analytes for up to a week, at a temperature of 2-8 (various studies), and in the case of the study, the gel preserved the triglyceride value while the glucose value did not, and this is related to the effect of glycolysis because this is a basic pro-
of the decrease in glucose concentration as a result of this phenomenon is slower as the temperature decreases. in this study, it was observed that the separating gel of the tube does not inhibit the process of glycolysis and as a result, the glycemic values changed. The change in blood glucose values may also have come as a result of the presence of blood elements in the serum. (number of leukocytes always interfere in this avoidance of glucose concentration values).
The second discussion is related to the use of a tube with sodium fluoride, to evaluate if, under the same analysis conditions, these obtained statistical relationships are preserved, which will provide valuable information regarding the obtained values (of blood, serum) and analytes).
Acknowledgement cial thanks goes to this institution as well as to my
This study was conducted in the Clinical Bio- colleagues. For the help, support and the review
chemistry Laboratory at Aldent University. A spe- carried out.
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