A Nanomaterial-Based Aptamer Fluorescence Sensing Strategy For Rapid Detection Of Staphylococcus Aureus Enterotoxin A

This work was supported by the National Natural Science Foundation of China(No.31571919).Food safety is one of the major public health problems in the world,especially the harm of food-borne pathogenic bacteria.Among them,food poisoning caused by enterotoxins produced by Staphylococcus aureus are more common.For the more than twenty Staphylococcus aureus enterotoxins(SEs),the proportion of food poisoning caused by Staphylococcus aureus enterotoxin A(SEA)is the highest.Chromatography and immunological analysis have the advantages of high efficiency and sensitivity.But,its development in the field of rapid detection is limited by the disadvantages of long time,complex operation and high cost.Aptamer is an emerging chemical antibody.Due to its small molecular weight,high stability,strong affinity,wide application and low cost,it has become an important identification element.At the same time,because the combination of nanomaterials and aptamers can improve the sensitivity of the detection,it has been widely developed.In this paper,the specific recognition of aptamers on target was combined with nanomaterials.Two simple and rapid fluorescence sensing strategies for efficient and sensitive detection of SEA were developed.The main contents of the study include:(1)Fluorophore-labeled aptamers were used as a molecular recognition element.The positively-charged gold nanorods bound to the nucleic acid through electrostatic adsorption,and the gold nanorods quenched the fluorescence.Then,a highly specific and sensitive SEA detection method was established.In the presence of SEA,SEA specifically bound to the aptamers,and gold nanorods was dissociated in solution,which reduced the FRET efficiency between the fluorophore and gold nanorods,resulting in the fluorescence was high.On the contrary,when there was no SEA,the gold nanorods were combined with fluorophore-labeled aptamers,and the fluorescence of the system was low.Under the optimal conditions,the SEA concentration in 0.01-0.8 ?g/mL was linearly correlated with the fluorescence intensity,and the detection limit was 5.66 ng/mL.This method has been successfully applied to SEA detection in milk without complicated pretreatment processes.It provided a simple and fast fluorescence detection method for SEA.(2)A rapid and sensitive method for SEA detection was developed based on unlabeled aptamers.The affinity of graphene oxide for single-stranded DNA and the characteristic that SYBR Green I specifically binds to double-stranded DNA were exploited.In this system,compared with the combination of aptamer and SEA,the affinity between the aptamer and its complementary strand was stronger.In the absence of SEA,aptamers bound to the graphene oxide,the compounds were separated from the solution by centrifugation.Then,complementary strand and SGI were added,and a weak fluorescence signals were generated.In the presence of SEA,the aptamer specifically bound to the SEA,the dissociated aptamer bound to the graphene oxide and was separated from the solution by centrifugation.Then,the complementary strand bound to the aptamer because of the high affinity,and formed double-stranded DNA.The SGI combined with double-stranded DNA to produce stronger fluorescence.Under the optimal conditions,there was a good linear relationship between SEA and fluorescence intensity in the range of 0.001-0.06 ?g/mL,and the detection limit was 0.667 ng/mL.This method avoided the influence of the marker on the affinity because the unlabeled aptamer.At the same time,the development of the method can be applied to the other targets without being limited by the length of the aptamer,so the method was more developable.