Limits...
Development of chemiluminescent lateral flow assay for the detection of nucleic acids.

Wang Y, Fill C, Nugen SR - Biosensors (Basel) (2012)

Bottom Line: Rapid, sensitive detection methods are of utmost importance for the identification of pathogens related to health and safety.On-membrane enzymatic signal amplification is used to reduce the limit of detection to the sub-femtomol level.The limit of detection was determined to be 0.5 fmols of nucleic acid target.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science, University of Massachusetts, 102 Holdsworth Way, Amherst, MA 01003, USA. yuhongw@foodsci.umass.edu.

ABSTRACT
Rapid, sensitive detection methods are of utmost importance for the identification of pathogens related to health and safety. Herein we report the development of a nucleic acid sequence-based lateral flow assay which achieves a low limit of detection using chemiluminescence. On-membrane enzymatic signal amplification is used to reduce the limit of detection to the sub-femtomol level. To demonstrate this assay, we detected synthetic nucleic acid sequences representative of Trypanosoma mRNA, the causative agent for African sleeping sickness, with relevance in human and animal health in sub-Saharan Africa. The intensity of the chemiluminescent signal was evaluated by using a charge-coupled device as well as a microtiter plate reader. We demonstrated that our lateral flow chemiluminescent assay has a very low limit of detection and is easy to use. The limit of detection was determined to be 0.5 fmols of nucleic acid target.

No MeSH data available.


Related in: MedlinePlus

Chemiluminescence at both the test lines and controls lines were quantified using a microtiter plate reader over 60 min. At low target amounts such as this (0.1 fmol), the control line appears much stronger than the test line.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4263543&req=5

biosensors-02-00032-f004: Chemiluminescence at both the test lines and controls lines were quantified using a microtiter plate reader over 60 min. At low target amounts such as this (0.1 fmol), the control line appears much stronger than the test line.

Mentions: Similar results were obtained using the microtiter plate reader. This assay allowed for quantification of the signal over time. A typical result obtained using the multi-mode microtiter plate reader is shown in Figure 4. At low target amounts, the chemiluminescent signal generated at the control line was significantly stronger than that at the test line which is in agreement with the results from the CCD imaging. The chemiluminescence generated on the membrane was detectable for over an hour following the application of the luminol-H2O2 mixture. The results suggested an increase in the signal generation with increasing target levels. Generally, the maximum chemiluminescent signal intensity of the target samples occurred at 20–30 min after application of the mixture, and the maximum intensity of negative control was less than 40 min. To quantify the signal, the signal intensity was integrated over a sixty minute period. The integration of the test line signal increased accordingly with increasing target concentration.


Development of chemiluminescent lateral flow assay for the detection of nucleic acids.

Wang Y, Fill C, Nugen SR - Biosensors (Basel) (2012)

Chemiluminescence at both the test lines and controls lines were quantified using a microtiter plate reader over 60 min. At low target amounts such as this (0.1 fmol), the control line appears much stronger than the test line.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263543&req=5

biosensors-02-00032-f004: Chemiluminescence at both the test lines and controls lines were quantified using a microtiter plate reader over 60 min. At low target amounts such as this (0.1 fmol), the control line appears much stronger than the test line.
Mentions: Similar results were obtained using the microtiter plate reader. This assay allowed for quantification of the signal over time. A typical result obtained using the multi-mode microtiter plate reader is shown in Figure 4. At low target amounts, the chemiluminescent signal generated at the control line was significantly stronger than that at the test line which is in agreement with the results from the CCD imaging. The chemiluminescence generated on the membrane was detectable for over an hour following the application of the luminol-H2O2 mixture. The results suggested an increase in the signal generation with increasing target levels. Generally, the maximum chemiluminescent signal intensity of the target samples occurred at 20–30 min after application of the mixture, and the maximum intensity of negative control was less than 40 min. To quantify the signal, the signal intensity was integrated over a sixty minute period. The integration of the test line signal increased accordingly with increasing target concentration.

Bottom Line: Rapid, sensitive detection methods are of utmost importance for the identification of pathogens related to health and safety.On-membrane enzymatic signal amplification is used to reduce the limit of detection to the sub-femtomol level.The limit of detection was determined to be 0.5 fmols of nucleic acid target.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science, University of Massachusetts, 102 Holdsworth Way, Amherst, MA 01003, USA. yuhongw@foodsci.umass.edu.

ABSTRACT
Rapid, sensitive detection methods are of utmost importance for the identification of pathogens related to health and safety. Herein we report the development of a nucleic acid sequence-based lateral flow assay which achieves a low limit of detection using chemiluminescence. On-membrane enzymatic signal amplification is used to reduce the limit of detection to the sub-femtomol level. To demonstrate this assay, we detected synthetic nucleic acid sequences representative of Trypanosoma mRNA, the causative agent for African sleeping sickness, with relevance in human and animal health in sub-Saharan Africa. The intensity of the chemiluminescent signal was evaluated by using a charge-coupled device as well as a microtiter plate reader. We demonstrated that our lateral flow chemiluminescent assay has a very low limit of detection and is easy to use. The limit of detection was determined to be 0.5 fmols of nucleic acid target.

No MeSH data available.


Related in: MedlinePlus