In vitro selection of DNA aptamers binding pesticide fluoroacetamide.

Fluoroacetamide (Mw = 77.06) is a lethal rodenticide to humans and animals which is still frequently abused in food storage somewhere in China. The production of antibodies for fluoroacetamide is difficult due to its high toxicity to animals, which limits the application of immunoassay method in poison detection. In this work, aptamers targeting N-fluoroacetyl glycine as an analog of fluoroacetamide were selected by a specific systematic evolution of ligands by exponential enrichment (SELEX) strategy. The binding ability of the selected aptamers to fluoroacetamide was identified using surface plasmon resonance (SPR)-based assay. The estimated KD values in the low micromolar range showed a good affinity of these aptamers to the target. Our work verified that the SELEX strategy has the potential for developing aptamers targeted to small molecular toxicants and aptamers can be employed as new recognition elements instead of antibodies for poison detection.

Development of highly sensitive and specific mRNA multiplex system (XCYR1) for forensic human body fluids and tissues identification.

The identification of human body fluids or tissues through mRNA-based profiling is very useful for forensic investigations. Previous studies have shown mRNA biomarkers are effective to identify the origin of biological samples. In this study, we selected 16 tissue specific biomarkers to evaluate their specificities and sensitivities for human body fluids and tissues identification, including porphobilinogen deaminase (PBGD), hemoglobin beta (HBB) and Glycophorin A (GLY) for circulatory blood, protamine 2 (PRM2) and transglutaminase 4 (TGM4) for semen, mucin 4 (MUC4) and human beta defensin 1(HBD1) for vaginal secretion, matrix metalloproteinases 7 and 11 (MMP7 and MMP11) for menstrual blood, keratin 4(KRT4) for oral mucosa, loricrin (LOR) and cystatin 6 (CST6) for skin, histatin 3(HTN3) for saliva, statherin (STATH) for nasal secretion, dermcidin (DCD) for sweat and uromodulin (UMOD) for urine. The above mentioned ten common forensic body fluids or tissues were used in the evaluation. Based on the evaluation, a reverse transcription (RT) PCR multiplex assay, XCYR1, which includes 12 biomarkers (i.e., HBB, GLY, HTN3, PRM2, KRT4, MMP11, MUC4, DCD, UMOD, MMP7, TGM4, and STATH) and 2 housekeeping genes [i.e., glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 18SrRNA], was developed. This assay was further validated with real casework samples and mock samples (with both single source and mixture) and it was approved that XCYR1 is effective to identify common body fluids or tissues (i.e., circulatory blood, saliva, semen, vaginal secretion, menstrual blood, oral mucosa, nasal secretion, sweat and urine) in forensic casework samples.

[Determination of bloodstain formation time by RNA analysis].

OBJECTIVE: To investigate the expression of 18S rRNA and beta-actin mRNA in bloodstain between 8 and 15 days after death and extrapolate the time of bloodstain formation. METHODS: RNA in dried bloodstain at different times was extracted, then quantified for 18S rRNA and beta-actin mRNA by real-time RT-PCR. The bloodstain formation time was deduced based on the changes of the ratio of 18S rRNA to beta-actin mRNA at different time points. RESULTS: The ratio of 18S rRNA to beta-actin mRNA increased gradually with time, indicating that rRNA and mRNA degraded in different rate with time. CONCLUSION; The ratio of 18S rRNA to beta-actin mRNA could be used for estimating the time of bloodstain formation in some period.

[Forensic analysis of LCN DNA using sample concentration methods followed by miniSTR genotyping].

OBJECTIVE: To optimize low copy number (LCN) DNA analysis methods for forensic STR genotyping. METHODS: Two groups of DNA sample, extracted using either Magnetic bead method or Chelex-100 methods, were previously amplified with a Identifiler PCR Amplification kit, but no genotype was detected. The DNA samples were concentrated using either a drying method or the Microcon-100 method, then amplified using an miniFiler PCR Amplification kit and genotyped. RESULTS: Among the 127 DNA samples, 47 samples, previously extracted using the Magnetic bead method, were genotyped with 36% success rate. Eighty samples, previously extracted using the Chelex-100 method, were genotyped with 30% success rate. CONCLUSION: The application of sample concentration methods and miniFiler kit can improve the success rate of LCN STR analysis.

A low power wearable transceiver for human body communication.

This paper reports a low power transceiver designed for wearable medical healthcare system. Based on a novel energy-efficient wideband wireless communication scheme that uses human body as a transmission medium, the transceiver can achieve a maximum 15 Mbps data rate with total receiver sensitivity of -30 dBm. The chip measures only 0.56 mm(2) and was fabricated in the SMIC 0.18um 1P6M RF CMOS process. The RX consumes 5mW and TX dissipates 1mW with delivering power up to 10uW, which is suitable for the body area network short range application. Real-time medical information collecting through the human body is fully simulated. Architecture of the chip together with the detail characterizes from its wireless analog front-end are presented.

[Study on colloidal gold labeled anti-buprenorphine monoclonal antibody for rapid test kit to detect buprenorphine].

OBJECTIVE: To develope an easy to use, rapid and accurate test for detecting buprenorphine based on the principle of competitive immunoassay. METHODS: Monoclonal antibody against buprenorphine was conjugated with colloidal gold and dispensed on the glass fiber. The complete antigen Buprenorphine-BSA and the goat anti-mouse IgG polyclonal antibody were separately sprayed on the nitrocellulose membrane as the test line (T line) and the control line (C line). The rapid test kit was the final assembled product of test strip with the plastic cover. RESULTS: A total of 100 urine samples were tested for buprenorphine by immunochromatographic and GC/ MS methods. The accuracy was 99.0%. It is found the test kit can only detect by cross reaction with other 45 kind drugs. CONCLUSION: Rapid test kit can detect buprenorphine in the samples in 5 minutes. The cut-off value of the test is 100 ng/mL.

[DNA genotyping of oral epithelial cells by laser capture microdissection].

OBJECTIVE: The STR genotypping of trace oral epithelial cells which are microdissected by laser capture microdissection system (LCM) is explored. METHODS: The oral epithelial cells are microdissected using a low-power infrared laser by VERITAS Microdissection Instrument. STR loci of Profiler Plus are detected by multiplex PCR procesures. RESULTS: DNA genotyping of 7-8 oral epithelial cells are succeeded, and DNA genotyping of 3-4 oral epithelial cells are failed. CONCLUSION: It is viable in genotyping of trace oral epithelial cells by Laser Capture Microdissection as a new technology of seperating single cell.