Laser-induced fluorescence integrated in a microfluidic immunosensor for quantification of human serum IgG antibodies to Helicobacter pylori

MARCO A. SEIA; PEREIRA SIRLEY V.; FONTAN CARLOS A.; IRMA E. DE VITO; GERMÁN A. MESSINA; JULIO RABA

SENSORS AND ACTUATORS B-CHEMICAL

ELSEVIER SCIENCE SA

Lugar: Amsterdam; Año: 2012

0925-4005

A portable immunosensor coupled to laser-induced fluorescence (LIF) detection  system has been successfully developed. It was applied to the quantitative determination  of specific IgG antibodies against H. pylori present in human serum samples. This  system is a relevant alternative tool for the diagnostic of peptic ulcer and gastritis  diseases produced by H. pylori, which are recognized risk factors for the development  of gastric mucosa associated lymphoid tissue lymphoma and gastric adenocarcinoma.  More than 50% of the population has been affected by H. pylori, one of the most  widespread infections around the world.  Our proposed system combines the LIF detection, which offers great sensibility  with the specificity of the immunological reactions and the microfluidic technology.  The device has a central channel (CC) with packed H. pylori antigen immobilized on 3- aminopropyl-modified controlled pore glass (AP-CPG). Antibodies in serum samples  reacted immunologically with the immobilized antigen and then, they were determined  using alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human  IgG. The 4-methylumbelliferyl phosphate (4-MUP), employed as enzymatic substrate,  was converted to soluble fluorescent methylumbelliferone by AP, and this fluorescent  product was finally quantified by LIF detection system. The calculated detection limits  for LIF detection and the ELISA procedure were 0.17 and 2.1 UmL−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.H. pylori present in human serum samples. This  system is a relevant alternative tool for the diagnostic of peptic ulcer and gastritis  diseases produced by H. pylori, which are recognized risk factors for the development  of gastric mucosa associated lymphoid tissue lymphoma and gastric adenocarcinoma.  More than 50% of the population has been affected by H. pylori, one of the most  widespread infections around the world.  Our proposed system combines the LIF detection, which offers great sensibility  with the specificity of the immunological reactions and the microfluidic technology.  The device has a central channel (CC) with packed H. pylori antigen immobilized on 3- aminopropyl-modified controlled pore glass (AP-CPG). Antibodies in serum samples  reacted immunologically with the immobilized antigen and then, they were determined  using alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human  IgG. The 4-methylumbelliferyl phosphate (4-MUP), employed as enzymatic substrate,  was converted to soluble fluorescent methylumbelliferone by AP, and this fluorescent  product was finally quantified by LIF detection system. The calculated detection limits  for LIF detection and the ELISA procedure were 0.17 and 2.1 UmL−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.H. pylori, which are recognized risk factors for the development  of gastric mucosa associated lymphoid tissue lymphoma and gastric adenocarcinoma.  More than 50% of the population has been affected by H. pylori, one of the most  widespread infections around the world.  Our proposed system combines the LIF detection, which offers great sensibility  with the specificity of the immunological reactions and the microfluidic technology.  The device has a central channel (CC) with packed H. pylori antigen immobilized on 3- aminopropyl-modified controlled pore glass (AP-CPG). Antibodies in serum samples  reacted immunologically with the immobilized antigen and then, they were determined  using alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human  IgG. The 4-methylumbelliferyl phosphate (4-MUP), employed as enzymatic substrate,  was converted to soluble fluorescent methylumbelliferone by AP, and this fluorescent  product was finally quantified by LIF detection system. The calculated detection limits  for LIF detection and the ELISA procedure were 0.17 and 2.1 UmL−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.H. pylori, one of the most  widespread infections around the world.  Our proposed system combines the LIF detection, which offers great sensibility  with the specificity of the immunological reactions and the microfluidic technology.  The device has a central channel (CC) with packed H. pylori antigen immobilized on 3- aminopropyl-modified controlled pore glass (AP-CPG). Antibodies in serum samples  reacted immunologically with the immobilized antigen and then, they were determined  using alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human  IgG. The 4-methylumbelliferyl phosphate (4-MUP), employed as enzymatic substrate,  was converted to soluble fluorescent methylumbelliferone by AP, and this fluorescent  product was finally quantified by LIF detection system. The calculated detection limits  for LIF detection and the ELISA procedure were 0.17 and 2.1 UmL−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.H. pylori antigen immobilized on 3- aminopropyl-modified controlled pore glass (AP-CPG). Antibodies in serum samples  reacted immunologically with the immobilized antigen and then, they were determined  using alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human  IgG. The 4-methylumbelliferyl phosphate (4-MUP), employed as enzymatic substrate,  was converted to soluble fluorescent methylumbelliferone by AP, and this fluorescent  product was finally quantified by LIF detection system. The calculated detection limits  for LIF detection and the ELISA procedure were 0.17 and 2.1 UmL−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.−1, respectively, and the within- and between-assay coefficients of variation were below 5.1%.