Research On Two Novel Technologies For Toxin Detection

Biotoxins are a class of chemically active substances produced by organisms such as animals,plants and microorganisms.Biotoxins can exhibit high toxicity,teratogenicity,and carcinogenicity to humans and animals,which seriously threaten human life and health.At the same time,biotoxins are characterized by wide sources,easy preparation,and easy transportation,and can be used as potential biological warfare agents and bioterrorism agents in terrorist incidents that endanger national security,posing a great threat to the security and stability of human society.In public health emergency diagnosis and treatment,when biotoxin-based poisoning occurs,due to the lack of effective clinical diagnosis methods and corresponding special antidote drugs,it is often difficult for poisoned patients to get timely and effective treatment and endanger their health and even their lives.The establishment of accurate and efficient toxin detection methods for different forms of biotoxin samples is of great significance for the clinical aid diagnosis of biotoxin poisoning.In this study,botulinum neurotoxin serotype A(Bo NT/A),ricin toxin(RT),and abrin toxin(AT),which are more toxic among biotoxins,were selected as the research objects.This study explored the application value of two new detection technologies,exhaled breath detection and nano-forest microfluidic chip,in the detection of biotoxins.In this study,a technology for the detection of botulism mice based on exhaled breath analysis and a method for the detection of ricin and abrin toxins based on nano-forest microfluidic chips were preliminarily established.The specific study includes the following two parts.In the first part,a model of Bo NT/A toxicity for mice was established by gas chromatography-mass spectrometry(GC-MS)using exhaled breath volatile organic compound(VOC)detection technique.The different components of exhaled breath from healthy mice and Bo NT/A toxicity mice were screened as markers of Bo NT/A toxicity.In the experiment,the stability of the Bo NT/A toxicity markers was verified by three different poisoning ways,intraperitoneal injection,gavage and pulmonary delivery.Moreover,three conditions including two neurotoxins poisoning and hypoxia were applied as controls to verify the specificity of the markers.This study demonstrates that three different intoxication methods significantly alter the levels of five markers,including acetone,ethanol,2-pentanone,n-butanol,and styrene,in the exhaled breath of healthy mice.Especially,the five markers did not produce the same changes in ricin poisoning,tetrodotoxin poisoning and hypoxia,indicating the stability and specificity of5 markers.In addition,through the tests of gas sensitivity,four metal oxides with good response to the 5 marker gases were selected from 10 metal oxide materials as sensitive electrodes.After optimization,three gas detection sensors with good detection effect and strong stability for the exhalation of mice with botulism were prepared using nano indium oxide,nano tin oxide,and iron oxide as sensitive materials,respectively.The above results lay a good foundation for further optimizing the detection of exhaled gas from botulism patients and developing electrochemical sensors for rapid detection of exhaled gas from botulism.In the second part,a self-driven microfluidic chip with nanoforest structure,capillary self-driving function,and super large surface area was designed by combining plasma repolymerization technology and double-antibody sandwich method in immunology.Specifically,one kind of nanoforest microfluidic detection chip was designed to detect RT in food samples and the other kind of microfluidic chip was designed to detect AT in food samples,which are produced by modifying different antibody molecules on the surface of the chips.These two microfluidic chips significantly outperform traditional immunochromatographic strips in terms of color signal uniformity and improve the sensitivity of both toxin detection,with the limit of detection of less than 10 pg/m L,while maintaining a good linear relationship in the range of 10 to 6250 pg/m L.Microfluidic chips can also be used to quickly detect RT or AT in simulated samples of fruit juice and milk,with detection sensitivity reaching 10pg/m L.Moreover,these two microfluidic chips have a simple structure,convenient operation,and fast detection abitlity of toxins in 15 minutes,which satisfies on-site rapid detection of RT and AT.In summary,this study successfully established two novel detection methods for the detection of biotoxins.Firstly,the feasibility of exhaled breath detection of botulism was initially verified and a set of electrochemical sensors with good gas-sensitive characteristics was prepared for the poisoning markers,which laid foundation for the subsequent development of exhaled breath detection sensors for clinical diagnosis and treatment of botulism.Secondly,the nanoforest microfluidic detection chip designed in this study for toxin detection has high detection sensitivity and can rapidly detect ricin and abrin in food samples such as juice and milk,which has good practical application scenarios and high socio-economic value.