[Establishment and application of quantitative immunoassay method based on stable element labeling and inductively coupled plasma mass spectrometry].

Objective: To establish a quantitative immunoassay method based on stable element labeling and inductively coupled plasma mass spectrometry (ICP-MS) for the detection of serum amyloid A (SAA) and evaluate its performance. Methods: An immunoassay system based on sandwich method was established with magnetic bead as carrier and holmium (Ho) as element tags. The binding ratio of hydrophilic streptavidin magnetic beads and biotinylated antibody, the amount of elemental antibody, and the reaction time were optimized to choose the optimal reaction conditions. According to the documents of Clinical and Laboratory Standards Institute (CLSI), the analytical performance was evaluated, including the limit of blank (LOB), linearity, accuracy, specificity, imprecision and interference test. Finally, 82 SAA plasma samples were collected after the turbidimetric inhibition immunoassay, and the newly established method was used for detection. Moreover, the detection results of the two methods were analyzed by Pearson correlation analysis. Results: The optimal binding ratio of hydrophilic streptavidin and biotinylated antibody was 1∶0.15, the amount of Ho-labeled antibody was 3 µl and the incubation time of the two reaction steps was 40 min and 30 min, respectively. The LOB was 0.6 ng/ml. The linearity was good within the range of 0-1 200 µg/L (R2=0.998 9, P<0.001). The inter-batch precision of high-value samples and low-value samples was 9.42% and 7.95%, respectively, and the intra-batch precision was 14.56% and 13.56%, respectively. The recovery was 96.01%-104.76%. The cross-reaction rates with procalcitonin (PCT) and C-reactive protein (CRP) were 0.45% and 0.015%, respectively. When the concentration of triglyceride≤35.5 mg/L, bilirubin≤0.52 mg/L and hemoglobin≤2.4 g/L, the interference bias was less than 10%. The results of 82 SAA plasma samples were 12.65 (4.45, 59.03) mg/L by ICP-MS immunoassay and 18.23 (9.33, 68.72) mg/L by turbidimetric inhibition immunoassay, respectively. The newly established system was well correlated with turbidimetric inhibition immunoassay (R2=0.983, P<0.001). Conclusion: The quantitative immunoassay for SAA with Ho as marker established in this study has high precision, good accuracy, high specificity, and wide linear range, which can meet the clinical testing requirements.

[Association between expression of plasma miRNA and the risk of childhood acute lymphocytic leukemia].

Objective: To investigate the characteristics of distribution and expression profiles of plasma miRNA in childhood acute lymphocytic leukemia (cALL) patients; the association between cALL incidence risk and plasma miRNA levels; the feasibility of plasma miRNA serving as cALL diagnostic biomarker. Methods: A total of 111 pairs of newly diagnosed cALL patients and patients with fractures were collected from Shenzhen Children's Hospital, China, between January 2015 and November 2016. Age and sex of the cases and controls were 1∶ 1 matched and LNA(TM) miRNA microarray was performed using 4 pairs of cALL and controls selected from the sample population. The expression level of miRNA was validated by real time quantitative PCR. Conditional logistic regression analysis was applied to evaluate the association between miRNA expression levels and the incidence risk of cALL. The receiver operating characteristic curve (ROC) and reclassification analysis were conducted to assess the feasibility of miRNAs serving as biomarkers for cALL. Results: A total of 204 differentially expressed miRNA were screened out and let-7f-5p, miR-5100, miR-25-3p and miR-3654 were selected for validation identified according to the inclusion criteria. The expression levels of let-7f-5p, miR-5100 and miR-25-3p in the cALL patients were significantly lower than those of the controls (P<0.01). After adjusting for confounding factors, 3 miRNAs remained significantly associated with the risk of cALL (OR and 95%CI were 0.84 (0.76-0.92), 0.81 (0.73-0.90) and 0.81 (0.74-0.89), respectively. Results from both the ROC analysis and reclassification analysis showed that introduction of one or more miRNA to traditional risk factors improved the area under the curve (P<0.05) and provided additional values to diagnosis (P<0.01). Conclusion: The expression levels of let-7f-5p, miR-5100 and miR-25-3p were significantly associated with the incidence rate of cALL, and these miRNAs might serve as promising biomarkers for cALL.

Reflectance confocal microscopy for the evaluation of sensitive skin.

BACKGROUND: Nowadays, the diagnosis for sensitive skin relies on subjective assessment or on the combination of subjective and objective evaluation. No quantitative evaluation is available. It could be expected that confocal microscopy imaging could be of interest to better define the condition. METHODS: Total 166 healthy female subjects were recruited in this study. Firstly, all subjects completed the sensitive questionnaire. Then, the cutaneous structures were measured by the reflectance confocal microscopy (RCM) on the face and fossa cubitalia. The lactic acid sting test was conducted finally. According to the results of self-perception sensitive skin questionnaire and lactic acid stinging test to evaluate facial skin sensitivity the both positive subjects were regarded as sensitive skin group and both negative group as healthy control group. RESULT: The results of RCM indicating that the proportion of 'disarranged honeycomb pattern' and 'spongiform edema' in the sensitive group and healthy control group were statistically different (P < 0.05), respectively; The following report 'damaged dermal papilla rings' was not a distinctive pattern, with no significant statistical difference (P > 0.05). The epidermal thickness was 38.88 ± 6.81 µm, healthy control group was 40.31 ± 9.37 µm in, respectively, sensitive skin group and healthy control group, there was no significant statistical difference between the two groups (P > 0.05). The honeycomb structure depth of sensitive group was 20.57 ± 4.86 µm. It was for 23.27 ± 6.38 µm, healthy control group the difference being statistically different between the two groups (P < 0.05). CONCLUSION: Based on the RCM results, 'epidermal honeycomb structure' and 'spongiform edema' may be used as new skin signs of RCM evaluation of sensitive skin effectively. Indeed, sensitive skin honeycomb structure depth was thinner compared with healthy control group. Such a specific pattern has good clinical and monitoring value for the further exploration. RCM could provide new data and patterns for the evaluation of sensitive skin.

[Association of exposure to environmental chemicals with risk of childhood acute lymphocytic leukemia].

Objective: This study aimed to investigate the association between exposure to environmental chemicals and the risk of childhood acute lymphocytic leukemia(cALL). Methods: A case-controlled study was conducted in Shenzhen Children's Hospital, China from January 2015 to January 2016. The cases were selected from the section of Hematology and Oncology, and the controls were selected from Orthopedics by 1∶2 matching of cases according to sex and age. A questionnaire including population data and chemical exposure characteristics was conducted on the children's parents, and urine and EDTA-blood were collected from the children. Then, we quantitatively measured the internal dose of formaldehyde(i.e., formaldehyde-human serum albumin)by enzyme-linked immunosorbent assay and the doses of metabolites benzene, toluene, and xylene(i.e., trans-muconic acid, hippuric acid, and methylhippuric acid)by high-performance liquid chromatography. Logistic regression models were used to analyze the relationships between exposure factors measured from children and their parents and cALL. Results: In the study, 71 cases(average age: 6.08±3.61 years), and 142 controls(average age: 5.91±3.57 years)were assessed; there were no differences in general demographics between two groups. The self-reported results showed that living in a home that had been painted in the past 10 years(OR=4.39, 95% CI: 1.87-10.31), maternal chemical exposure during pregnancy(OR=11.78, 95% CI: 1.65-83.88), paternal diesel or gasoline exposure(OR=8.15, 95% CI: 2.68-24.83), paternal dye exposure(OR=7.77, 95% CI: 1.52-39.67)and trash burning near the child's residence(OR=6.08, 95% CI: 1.17-31.66)were associated with increased risk of cALL. The positive detection rates of only benzene metabolites were significantly higher in cases(40/44)than controls(81/111)(χ2=5.92, P=0.021). The median formaldehyde and benzene concentrations in cases(32.120 pg/ml, 2.505 µg/gCr)were significantly higher than those in controls(18.705 pg/ml, 0.672 µg/gCr; Z values:-1.98 and-3.95, P values: 0.047 and<0.001, respectively). Multiple logistic regression analysis showed that benzene exposure(OR=1.09, 95% CI: 1.00-1.19), home painting in the past 10 years(OR=3.56, 95% CI: 1.20-10.53)and paternal diesel or gasoline exposure(OR= 3.75, 95% CI: 1.06-13.22)were associated with increased risk of cALL. Conclusion: A variety of environmental chemistry factors, such as benzene exposure, increase the risk of cALL, and further studies are warranted to explore their specific roles.