Development of a method for the determination of the JAK2 gene mRNA in venous blood and assessment of its diagnostic value in oncohematology

2021 ◽  
Vol 66 (6) ◽  
pp. 379-384
Author(s):  
M. A. Stolyar ◽  
A. S. Gorbenko ◽  
I. A. Olkhovskiy ◽  
V. I. Bakhtina ◽  
M. A. Mikhalev ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Venous Blood ◽  
Diagnostic Value ◽  
Janus Kinase ◽  
Rt Pcr ◽  
Blood Samples ◽  
Dnase Treatment ◽  
Pcr Method ◽  
Gene Mrna

Overactive JAK pathway signaling is a hallmark of immune diseases and critically affects on inflammation and coagulation. A number of mutations in the JAK2 gene act as driving forces of myeloproliferative neoplasms (MPN), the pathogenesis of certain variants of acute leukemia, a number of solid malignancies and cardiovascular diseases. Assays for quantifying JAK2 mRNA in circulating blood cells can be used as a marker associated with the activity of this enzyme. Development of an original method for detecting JAK2 mRNA in venous blood and assessment of the possible diagnostic value in chronic oncohematological diseases. The development of an RT-PCR method for determining the expression of the JAK2 gene mRNA in venous blood samples was carried out in accordance with the MIQE requirements. Primers and TaqMan probes were designed using the Primer3 program, taking into account the possibility of excluding subsequent DNase treatment. The stability of the investigated mRNA was assessed in vacutainers with different anticoagulants and depending on the storage time of the samples. The study of the expression of JAK2 mRNA in blood leukocytes of 41 patients with B-CLL, 16 patients with CML, 12 patients with multiple myeloma and 39 donors using the developed “real-time” PCR method. The study revealed a decrease in the level of JAK2 mRNA in venous blood samples in patients with primary CLL, but not with CML or with multiple myeloma. The level of the marker in the majority of patients with CLL after the start of therapy returned to the range typical for healthy people. It has been shown that the values of the relative expression of JAK2 mRNA are most stable in the range of 2 - 7 hours after taking blood in a vacutainer with EDTA. An original RT-PCR method was developed for the quantitative determination of JAK2 mRNA in venous blood samples, which meets the requirements of the MIQE system. Determination of JAK2 mRNA can be useful for clarifying the pathogenesis features of certain diseases involving impaired Janus kinase activity and can become a promising marker for prognosis and assessment of the effectiveness of therapy.

Developing method for allelic somatic mutations burden of Janus kinase 2 (V617F JAK2) detection in pools of venous blood samples

2016 ◽  
Vol 5 (1) ◽  
pp. 19 ◽  
Author(s):  
A. S. Gorbenko ◽  
M. A. Stolyar ◽  
T. N. Subbotina ◽  
M. A. Mihalev ◽  
I. A. Olkhovskiy
Keyword(s):  
Somatic Mutations ◽  
Venous Blood ◽  
Janus Kinase ◽  
Janus Kinase 2 ◽  
Blood Samples ◽  
Developing Method

scholarly journals PENENTUAN JENIS KELAMIN BURUNG KENARI (Serinus canaria) BERDASARKAN GEN Chromodomain Helicase DNA-Binding 1 (CHD1)

2020 ◽  
Vol 7 (1) ◽  
Author(s):  
Afif Muhammad Akhrom ◽  
Indarjulianto Soedarmanto ◽  
Yanuartono Yanuartono ◽  
Trini Susmiati ◽  
Alfarisa Nururrozi ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Sex Determination ◽  
Mature Female ◽  
Mature Male ◽  
Chain Reaction ◽  
Blood Samples ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Female Canaries

Phenotype determination of sex in young canaries is very low in accuracy. This study aimed to develop a genotypic sexing method in canaries. This study used 12 canaries consisting of 3 mature males, 3 mature females and 6 one-month-old canaries. Phenotypic sexing by cloacal observation was done on all birds, continued by genotypic sexing to identification CHD1 gene using polymerase chain reaction (PCR). The PCR used blood samples for mature canaries, and feather for mature and one-month-old canaries. The results of phenotypic observations showed that all mature male canaries had prominent and pointed cloaca forms, all mature females had flat and wide, whereas all one-month-old birds had a flat cloaca. The result of PCR showed a single band (500 bp) for mature male and double bands (500 bp and 300 bp) for mature female canaries. The PCR results of one-month-old canaries showed that there were one male and five females. Based on this study, it was concluded that genotypic sexing using the PCR method is effective in the sex determination of canaries.Keywords: canary, CHD1, genotype, PCR, sexing ABSTRAKPenentuan jenis kelamin burung kenari muda secara fenotip akurasinya sangat rendah. Penelitian ini bertujuan untuk menentukan jenis kelamin burung kenari secara genotip. Penelitian ini menggunakan 12 ekor burung kenari, terdiri dari 6 ekor dewasa (3 jantan, 3 betina) serta 6 ekor umur 1 bulan. Semua burung ditentukan jenis kelaminnya dengan mengamati kloaka dan identifikasi gen CHD1 menggunakan teknik polymerase chain reaction (PCR). Sampel DNA berasal dari darah dan bulu untuk burung dewasa serta bulu untuk burung umur 1 bulan. Pengamatan fenotip menunjukkan bahwa burung kenari dewasa jantan mempunyai bentuk kloaka menonjol dan runcing, dewasa betina berbentuk datar dan lebar, sedangkan semua burung umur 1 bulan mempunya bentuk kloaka datar. Hasil identifikasi gen CHD1 diperoleh adanya 1 pita gen sekitar 500 bp dari sampel darah dan bulu semua burung kenari dewasa jantan, dan 2 pita gen sekitar 500 bp dan 300 bp dari sampel semua burung kenari betina dewasa. Hasil PCR pada sampel burung umur 1 bulan menunjukkan bahwa 1 ekor jantan dan 5 ekor betina. Berdasarkan penelitian ini dapat disimpulkan bahwa penentuan jenis kelamin secara genotip menggunakan gen CHD1 dapat dilakukan pada burung kenari.

scholarly journals Sheep pestivirus in Morocco: sero-epidemiological and molecular study

2019 ◽  
Vol 6 (1) ◽  
pp. e000324 ◽  
Author(s):  
Ouafaa Fassi Fihri ◽  
Noâma Jammar ◽  
Nadia Amrani ◽  
Ikhlass El Berbri ◽  
Said Alali
Keyword(s):  
Reverse Transcriptase ◽  
Indirect Elisa ◽  
Molecular Study ◽  
Rt Pcr ◽  
Blood Samples ◽  
Sheep Breeding ◽  
Border Disease ◽  
Elisa Technique ◽  
The Difference

The present study is the first to investigate Border disease caused by the sheep pestivirus (SPV) in sheep herds in Morocco. Sero-epidemiological investigations were carried out in six regions of the Kingdom, known as important in terms of sheep breeding. A total of 760 blood samples were collected including aborted ewes from 28 randomly selected farms. The samples were analysed, for the determination of anti-pestivirus antibodies, using indirect ELISA technique. Next, reverse transcriptase PCR (RT-PCR) was conducted on serologically negative samples to identify possible persistently infected (PI) animals, through detection of specific RNA fragment. The results revealed an overall SPV seroprevalence in studied areas of 28.9%. The difference in seroprevalence between the six investigated regions was not statistically significant (p>0.05) and varied slightly from 20.9% to 37.5%. Furthermore, 93% of investigated farms were affected with an average seroprevalence of 22.7% (with a variation of 1%–74%). RT-PCR results were all negative, indicating the absence of PI animals in the tested samples. Nevertheless, the present study revealed that SPV is endemic in Morocco.

scholarly journals Tumor quantification of several fluoropyrimidines resistance gene expression with a unique quantitative RT-PCR method. Implications for pretherapeutic determination of tumor resistance phenotype

2006 ◽  
Vol 242 (2) ◽  
pp. 168-179 ◽  
Author(s):  
Maud Barbado ◽  
Laurence Preisser ◽  
Michele Boisdron-Celle ◽  
Veronique Verriele ◽  
Gerard Lorimier ◽  
...  
Keyword(s):  
Gene Expression ◽  
Resistance Gene ◽  
Tumor Resistance ◽  
Rt Pcr ◽  
Resistance Phenotype ◽  
Pcr Method

scholarly journals A highly specific real-time RT-PCR method for the quantitative determination of CK-19 mRNA positive cells in peripheral blood of patients with operable breast cancer

2006 ◽  
Vol 119 (7) ◽  
pp. 1654-1659 ◽  
Author(s):  
Aliki Stathopoulou ◽  
Maria Ntoulia ◽  
Maria Perraki ◽  
Stella Apostolaki ◽  
Dimitris Mavroudis ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Quantitative Determination ◽  
Real Time ◽  
Peripheral Blood ◽  
Operable Breast Cancer ◽  
Rt Pcr ◽  
Pcr Method

scholarly journals Crizotinib efficacy in advanced non-squamous NSCLC patients with ALK or ROS1 rearrangement

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Paweł Krawczyk ◽  
Anna Grenda ◽  
Paulina Terlecka ◽  
Justyna Błach ◽  
Kamila Wojas-Krawczyk ◽  
...  
Keyword(s):  
Predictive Factors ◽  
Targeted Therapies ◽  
Personalized Treatment ◽  
Genetic Diagnostics ◽  
Rt Pcr ◽  
Risk Of Death ◽  
Pcr Method ◽  
Nsclc Patients ◽  
L1 Protein

AbstractIn patients with advanced non-small cell lung cancer (NSCLC), comprehensive genetic diagnostics is currently carried out in order to qualify for molecularly targeted therapies and immunotherapy. The aim of the study was to assess the usefulness of the reverse transcriptase (RT-PCR) method in the diagnosis of gene rearrangements, the effectiveness of EGFR, ALK, ROS1, and PD-L1 inhibitors in first-line treatment in NSCLC patients. We enrolled 95 non-squamous NSCLC patients with known status of EGFR, ALK, ROS1, MET and RET genes and PD-L1 protein expression. We used the real time PCR, fluorescence in situ hybridization (FISH), immunohistochemistry (IHC) and RT-PCR techniques for determination of predictive factors. In patients with ALK and ROS1 genes alteration, the median overall survival was 34 months in crizotinib treated patients and 6 months in patients who received chemotherapy (HR = 0.266, p = 0.0056). The risk of death was lower in patients treated with molecularly targeted therapies or immunotherapy compared to patients with predictive factors without personalized treatment (HR = 0.265, 95% CI 0.116–0.606) and to patient without predictive factors who received chemotherapy (HR = 0.42, 95% CI 0.162–1.09). Diagnosis of predictive factors and implementation of personalized treatment are key to prolonging the survival in advanced NSCLC patients.

Vergleich von hämostaseologischen Parametern im arteriellen und venösen Blut bei Patienten mit peripherer arterieller Verschlußkrankheit

VASA ◽  
1999 ◽  
Vol 28 (1) ◽  
pp. 10-14 ◽  
Author(s):  
Woller ◽  
Lawall ◽  
Amann ◽  
Angelkort
Keyword(s):  
Venous Blood ◽  
Arterial System ◽  
Coagulation Activation ◽  
Blood Samples ◽  
Coagulation Parameters ◽  
Arterial Blood ◽  
Artery Disease ◽  
Peripheral Arterial ◽  
D Dimer

Background: Several studies proved the co-existence of peripheral arterial occlusive disease (PAOD) and hypercoagulability. However, in practice coagulation parameters are mainly determined from venous blood samples. In this study several coagulation parameters in arterial and venous blood were examined for differences and the validity of coagulation parameters determined in venous blood was investigated. Patients and methods: In 22 patients with peripheral artery disease venous and arterial blood samples from vessels of the diseased leg were examined for the concentration of thrombine-antithrombine III-complex (TAT), prothrombin fragments (F1 and F2) and D-dimers, and results were compared. Results: Mean concentrations of TATs and prothrombin fragments F1 and F2 were significantly higher in arterial than in venous blood. TAT-complex was the most sensitive parameter for quantification of thrombin generation. D-dimer levels did not differ in arterial and venous blood. TAT and F1 and F2 concentrations in arterial and venous blood did not correlate in individual patients whereas D-dimer concentration did. Conclusion: The determination of TAT and F1+F2 in venous blood does not adequately reflect the degree of the local coagulation activation in the arterial system. As indicators for hypercoagulability, D-Dimer values are less sensitive than F1+2, but venous D-dimer concentrations mirror arterial levels.

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