Resul Karakus

Effect of Blood Collection Tube Types on the Measurement of Human Epidermal Growth Factor

Abstract We observed significant differences in measured human epidermal growth factor (hEGF) levels for the same individuals serum/plasma samples between different tube types (glass, polystyrene, plastic with clot activator, plastic without clot activator, plastic with EDTA, polypropylene tubes). For all individuals, hEGF levels in plasma were found to be below the detection limit. The discrepancy of the hEGF levels in serum and plasma was attributed to the platelet derived EGF by analyzing platelet lyzate with size exclusion chromotography and demonstrating the immunoreactivity of the fractions corresponding to the pre‐proEGF and/or proEGF elution time. Besides, samples of females showed much higher EGF levels than those of males in certain test tube types. As a conclusion, all blood samples should be taken and stored in the same type of test tubes in order to make precise measurements for hEGF. And, the measured hEGF level in blood is susceptible to changes with blood clotting.

A New Efficient Method for Eliminating the Interference Effect of Human Serum and Increasing the Sensitivity and Recovery Rate of Enzyme Immunoassay

Abstract A new, very simple method for increasing the sensitivity and recovery rate of enzyme‐linked immunosorbent assay (ELISA) for the precise quantification of antigen in human serum is described. The assay design uses CATNF6A4c IgG2a monoclonal antibody and biotinylated anti‐human tumor necrosis factor‐α (hTNF‐α) polyclonal mouse IgG as the capture and tracer antibodies, respectively. The assay is completed within 4 hours at room temperature and is capable of detecting both recombinant and native human TNF‐α. The assay incorporates the use of saturated ammonium sulfate (SAS) as a component of the dilution buffer to amplify the resultant signal from antigen containing human serum and eliminating the endogenous interference of native human serum. SAS worked optimally at the final concentrations, ranging from 1.2% to 11%. The addition of SAS to the dilution buffer resulted in a dramatic increase in both sensitivity and recovery rate of the ELISA. The results demonstrated that 50 µL of dilution buffer, containing SAS, enabled the precise quantification of human TNF‐α in 100 µL of human serum samples and eliminated the interference of native serum, which seemed to be related to complement proteins. Therefore, dilution buffer containing SAS, at a defined concentration, seemed to be a potential candidate for resolving sensitivity and recovery problems usually encountered in immunoassays when measurement was performed with native serum samples. The proposed technique provides an easy, practical, and consistent method for ELISA when using human native serum.

Determinants of Protection against Diphtheria in Adult Hemodialysis Patients

Diphtheria is of great epidemiological concern. Although mainly observed during childhood, unvaccinated adults and relatively immunocompromised patients are at increased risk for acquiring diphtheria. We aimed to determine the rates and certain determinants of protection against diphtheria in adult hemodialysis (HD) patients. Protection rates of 322 HD patients were compared with 65 diabetes mellitus type 2 (DM) patients and 65 healthy controls. A questionnaire was held in regard to smoking habits and alcohol intake. Antibody levels against diphtheria were assessed by an in-house ELISA and a concentration of ≥0.1 IU/mL was regarded as protective. Effects of age, gender and time being on dialysis on protection were assessed by logistic regression. Ratios of individuals with protective antibody levels were found to be 36% (116/322), 27.7% (18/65), and 52.3% (34/65) for HD, DM, and control groups, respectively. Hemodialysis patients had a significantly (p < 0.05) lower protection rate than healthy controls. In all study groups, there was a tendency of higher protection rate with increasing age. These low ratios of protected individuals in both HD and DM patient groups are alarming, as these patients generally have defects in vaccine responses, and carriage is important in the perpetuation of diphtheria. The protection status of these patient groups might be improved with additional vaccinations.

Anaplastic lymphoma kinase gene expression in small round cell tumors of childhood--a comparative ımmunohistochemical study.

The focus of this study was to investigate anaplastic lymphoma kinase (ALK) expression by immunohistochemistry using a highly specific antibody. Distribution and frequency of ALK expression may provide a clue for ALK inhibitor use in small round cell tumors of childhood. The study group involved 76 small round cell tumors of childhood, which composed of 11 rhabdomyosarcomas, 13 Wilms tumors, 7 Ewing sarcoma/primitive neuroectodermal tumors, 34 peripheral neuroblastic tumors, and 11 acute lymphoblastic lymphoma. Anaplastic lymphoma kinase protein expression in small round cell tumors of childhood is poorly described in the literature. The findings of our study highlight a potential and possible role of targeting ALK in pediatric solid tumors by using ALK immunohistochemistry. Anaplastic lymphoma kinase may also have an oncogenic role in rhabdomyosarcomas and peripheral neuroblastic tumors, and they may possibly be treated with ALK inhibitors. Anaplastic lymphoma kinase expression in Wilms tumors is not reported in the literature, previously. Our study evaluated ALK expression in Wilms tumor samples.

Expression of Epstein-Barr virus in children with sacrococcygeal pilonidal sinus determined by immunohistochemical methods.

In this study, we probed whether chronic infections of skin such as pilonidal sinus could be a potential site of Epstein–Barr virus (EBV) replication. Pilonidal sinus is associated with a high recurrence rate. Therefore, we decided to determine the role of EBVs presence to explain whether it is correlated with the recurrence of pilonidal sinuses. This study was conducted on 36 patient samples with sacrococcygeal pilonidal sinus. Samples were immunohistochemically stained for EBV, CD3 and CD20 expression. Thirty‐six adolescents with pilonidal disease were evaluated. EBV‐positive cells were located in dermis with high inflammatory activity. EBV‐positive cells stained positive for the B‐cell antigen CD20 and were detected in 10 of 36 (27%) pilonidal sinus specimens. Among those who had experienced a relapse, three were positive for EBV expression. In addition, EBV expression was detected in eight cases with severe inflammation, and in two with minimal or moderate inflammation. Our study advances the field by demonstrating that similar to gastrointestinal mucosa, skin could be a reservoir for EBV. EBV was found to be restricted to B cells in skin lesions, and it was found that skin lesions with severe inflammation showed higher frequency of EBV expression in comparison to minimal or moderately inflammed skin lesions. Additionally, recurrence was more frequently observed among EBV‐positive cases. These findings point out for a role of EBV infection in the recurrence of pilonidal sinuses.