• The Korean Journal of Nuclear Medicine Technology
• /
• v.20 no.2
• /
• pp.62-68
• /
• 2016

Purpose In a cyclosporine experiment using a robotic liquid handing system has found a deviation of its standard curve and low reproducibility of patients's results. The difference of the test is that methanol is mixed with samples and the extractions are used for the test. Therefore, we assumed that the abnormal test results came from using methanol and conducted this test. In a manual of a robotic liquid handling system mentions that we can choose several setting parameters depending on the viscosity of the liquids being used, the size of the sampling tips and the motor speeds that you elect to use but there's no exact order. This study was undertaken to confirm pipetting ability depending on types of liquids and investigate proper setting parameters for the optimum dispensing ability. Materials and Methods 4types of liquids(water, serum, methanol, PEG 6000(25%)) and $TSH^{125}I$ tracer(515 kBq) are used to confirm pipetting ability. 29 specimens for Cyclosporine test are used to compare results. Prepare 8 plastic tubes for each of the liquids and with multi pipette $400{\mu}l$ of each liquid is dispensed to 8 tubes and $100{\mu}l$ of $TSH^{125}I$ tracer are dispensed to all of the tubes. From the prepared samples, $100{\mu}l$ of liquids are dispensed using a robotic liquid handing system, counted and calculated its CV(%) depending on types of liquids. And then by adjusting several setting parameters(air gap, dispense time, delay time) the change of the CV(%)are calcutated and finds optimum setting parameters. 29 specimens are tested with 3 methods. The first(A) is manual method and the second(B) is used robotic liquid handling system with existing parameters. The third(C) is used robotic liquid handling system with adjusted parameters. Pipetting ability depending on types of liquids is assessed with CV(%). On the basis of (A), patients's test results are compared (A)and(B), (A)and(C) and they are assessed with %RE(%Relative error) and %Diff(%Difference). Results The CV(%) of the CPM depending on liquid types were water 0.88, serum 0.95, methanol 10.22 and PEG 0.68. As expected dispensing of methanol using a liquid handling system was the problem and others were good. The methanol's dispensing were conducted by adjusting several setting parameters. When transport air gap 0 was adjusted to 2 and 5, CV(%) were 20.16, 12.54 and when system air gap 0 was adjusted to 2 and 5, CV(%) were 8.94, 1.36. When adjusted to system air gap 2, transport air gap 2 was 12.96 and adjusted to system air gap 5, Transport air gap 5 was 1.33. When dispense speed was adjusted 300 to 100, CV(%) was 13.32 and when dispense delay was adjusted 200 to 100 was 13.55. When compared (B) to (A), the result increased 99.44% and %RE was 93.59%. When compared (C-system air gap was adjusted 0 to 5) to (A), the result increased 6.75% and %RE was 5.10%. Conclusion Adjusting speed and delay time of aspiration and dispense was meaningless but changing system air gap was effective. By adjusting several parameters proper value was found and it affected the practical result of the experiment. To optimize the system active efforts are needed through the test and in case of dispensing new types of liquids proper test is required to check the liquid is suitable for using the equipment.

• Tuberculosis and Respiratory Diseases
• /
• v.41 no.4
• /
• pp.339-353
• /
• 1994

Background : The p53 gene codes for a DNA-binding nuclear phosphoprotein that appears to inhibit the progression of cells from the G1 to the S phase of the cell cycle. Mutations of the p53 gene are common in a wide variety of human cancers, including lung cancer. In lung cancers, point mutations of the p53 gene have been found in all histological types including approximately 45% of resected NSCLC and even more frequently in SCLC specimens. Mutant forms of the p53 protein have transforming activity and interfere with the cell-cycle regulatory function of the wild-type protein. The majority of p53 gene mutations produce proteins with altered conformation and prolonged half life; these mutant proteins accumulate in the cell nucleus and can be detected by immunohistochemical staining. But protein overexpression has been reported in the absence of mutation. p53 protein overexpression or gene mutation is reported poor prognostic factor in breast cancer, but in lung cancer, its prognostic significance is controversial. Method : We investigated the p53 abnormalities by nucleotide sequencing, polymerase chain reaction-single strand conformation polymorphism(PCR-SSCP), and immunohistochemical staining. We correlated these results with each other and survival in 75 patients with NSCLC resected with curative intent. Overexpression of the p53 protein was studied immunohistochemically in archival paraffin- embedded tumor samples using the D07(Novocastra, U.K.) antibody. Overexpression of p53 protein was defined by the nuclear staining of greater than 25% immunopositive cells in tumors. Detection of p53 gene mutation was done by PCR-SSCP and nucleotide sequencing from the exon 5-9 of p53 gene. Result: 1) Of the 75 patients, 36%(27/75) showed p53 overexpression by immunohistochemical stain. There was no survival difference between positive and negative p53 immunostaining(overall median survival of 26 months, disease free median survival of 13 months in both groups). 2) By PCR-SSCP, 27.6%(16/58) of the patients showed mobility shift. There was no significant difference in survival according to mobility shift(overall median survival of 27 in patients without mobility shift vs 20 months in patients with mobility shift, disease free median survival of 8 months vs 10 months respectively). 3) Nucleotide sequence was analysed from 29 patients, and 34.5%(10/29) had mutant p53 sequence. Patients with the presence of gene mutations showed tendency to shortened survival compared with the patients with no mutation(overall median survival of 22 vs 27 months, disease free median survival of 10 vs 20 months), but there was no statistical significance. 4) The sensitivity and specificity of immunostain based on PCR-SSCP was 67.0%, 74.0%, and that of the PCR-SSCP based on the nucleotide sequencing was 91.8%, 96.2% respectively. The concordance rate between the immunostain and PCR-SSCP was 62.5%, and the rate between the PCR-SSCP and nucleotide sequencing was 95.3%. Conclusion : In terms of detection of p53 gene mutation, PCR-SSCP was superior to immunostaining. p53 gene abnormalities either overexpression or mutation were not a significant prognostic factor in NSCLC patients resected with curative intent. However, patients with the mutated p53 gene showed the trends of early relapse.