Researchers at Osaka Metropolitan University have developed an optical alternative to immunoassays and other methods used for protein analysis. The alternative method provides rapid, highly sensitive detection of proteins through laser irradiation.
According to the researchers, the light-induced acceleration-based technique could improve detection limit and quantitative measurement, using a small number of biological samples and a simple process, to aid in the ultra-early diagnosis of cancer, dementia, and infectious diseases.
Conventional techniques for protein detection, such as enzyme-linked immunosorbent assay (ELISA), require several hours and involve multiple steps, in addition to being less sensitive than the recently developed light-induced method.
In experiments, the researchers showed a successful deployment of their approach using only three minutes of laser irradiation. They achieved a sensitivity and ultrafast specific detection more than 100× that obtained in comparison with conventional protein detection methods. Further, the researchers showed that the technique could enable diagnoses with only a small amount of body fluids — such as a single drop of blood.
To develop an optical method to achieve control of antigen-antibody reaction and detect trace amounts of proteins, the researchers conducted basic research on the synergistic effects of optical pressure and fluid pressure and how to circumvent the effect of heat. They used target proteins, to which they introduced probe particles containing modified antibodies that selectively bound to the proteins. They confined the proteins and probe particles to a microchannel and irradiated the channel.
The probe particles were 2-μm-diameter polymer beads with a minimal amount of heat generation, due to the absorption of infrared laser light as well as strong light scattering.
The researchers then used light-induced acceleration to trap antigen-antibody reactions of trace amounts of proteins at the interface between solid and liquid (i.e., the bottom of the channel, which contained liquid samples).
After tuning the laser irradiation area to be comparable to the confinement geometry, the researchers irradiated a few hundred milliwatts of laser light, defocused to a spot size of approximately 70 μm in the microchannel, which had a width of approximately 100 μm.
The laser-assisted optical pressure on the proteins and probe particles increased the probability of interaction and the acceleration of antigen-antibody reactions. The collisional probability of the target molecules and probe particles was enhanced through optical force and fluidic pressure.
The “scattering force,” a component of the optical force, was enhanced to ensure accumulating force without any thermal damage to the antibody-modified probe particles and target proteins.
After testing various conditions, the researchers found that the antigen-antibody reaction was efficiently accelerated by adjusting the flow rate to 100 to 200 μm/s.
A black region formed in a portion of the assembled structure obtained by laser irradiation, because the optical transmission was blocked by the multilayered structure formation. The researchers found that the area of this region was positively correlated with the protein concentration. A model calculation, in which binding by an antigen-antibody reaction was expressed using cohesive energy, confirmed theoretically that the formation of the multilayered structure was caused by optical force and pressure-driven flow.
When the researchers irradiated the microchannel with IR laser light for three minutes, they were able to detect trace amounts of proteins at a sensitivity level approximately 100× higher than that of conventional protein testing. The researchers achieved rapid measurement of trace amounts on the order of tens of attograms (ag) (ag = 10−18 g; one quintillionth of a gram). They measured target protein trace amounts as small as one twenty-quadrillionth of a gram after only three minutes of irradiation.
The researchers applied the principle of light-induced acceleration to several different types of membrane proteins. In experiments, the optical technique demonstrated ultrafast, specific detection of target proteins with a smaller sample volume and higher sensitivity than conventional techniques. For example, in one type of membrane protein, the researchers detected 47 to 750 ag of target proteins, without any pretreatment, from a 300-nL sample after just three minutes of laser irradiation.
A progressive collaborative study on cancer marker measurement using patient-derived samples is underway as part of the Future Society Creation Project of the Japan Science and Technology Agency. An initial validation of the light-induced acceleration technique in clinical practice is planned, with the aim of developing a basic system within a few years, the researchers said.
Since antigen-antibody reaction is a common biochemical reaction, the technology has the potential to be used not only in the medical field but also in various industrial fields, such as testing for allergens in food and drink, detecting biological substances in the environment, and testing for intermediate products in the pharmaceutical process.
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According to the researchers, the light-induced acceleration-based technique could improve detection limit and quantitative measurement, using a small number of biological samples and a simple process, to aid in the ultra-early diagnosis of cancer, dementia, and infectious diseases.
Conventional techniques for protein detection, such as enzyme-linked immunosorbent assay (ELISA), require several hours and involve multiple steps, in addition to being less sensitive than the recently developed light-induced method.
In experiments, the researchers showed a successful deployment of their approach using only three minutes of laser irradiation. They achieved a sensitivity and ultrafast specific detection more than 100× that obtained in comparison with conventional protein detection methods. Further, the researchers showed that the technique could enable diagnoses with only a small amount of body fluids — such as a single drop of blood.
To develop an optical method to achieve control of antigen-antibody reaction and detect trace amounts of proteins, the researchers conducted basic research on the synergistic effects of optical pressure and fluid pressure and how to circumvent the effect of heat. They used target proteins, to which they introduced probe particles containing modified antibodies that selectively bound to the proteins. They confined the proteins and probe particles to a microchannel and irradiated the channel.
The probe particles were 2-μm-diameter polymer beads with a minimal amount of heat generation, due to the absorption of infrared laser light as well as strong light scattering.
The researchers then used light-induced acceleration to trap antigen-antibody reactions of trace amounts of proteins at the interface between solid and liquid (i.e., the bottom of the channel, which contained liquid samples).
After tuning the laser irradiation area to be comparable to the confinement geometry, the researchers irradiated a few hundred milliwatts of laser light, defocused to a spot size of approximately 70 μm in the microchannel, which had a width of approximately 100 μm.
The laser-assisted optical pressure on the proteins and probe particles increased the probability of interaction and the acceleration of antigen-antibody reactions. The collisional probability of the target molecules and probe particles was enhanced through optical force and fluidic pressure.
The “scattering force,” a component of the optical force, was enhanced to ensure accumulating force without any thermal damage to the antibody-modified probe particles and target proteins.
After testing various conditions, the researchers found that the antigen-antibody reaction was efficiently accelerated by adjusting the flow rate to 100 to 200 μm/s.
A black region formed in a portion of the assembled structure obtained by laser irradiation, because the optical transmission was blocked by the multilayered structure formation. The researchers found that the area of this region was positively correlated with the protein concentration. A model calculation, in which binding by an antigen-antibody reaction was expressed using cohesive energy, confirmed theoretically that the formation of the multilayered structure was caused by optical force and pressure-driven flow.
When the researchers irradiated the microchannel with IR laser light for three minutes, they were able to detect trace amounts of proteins at a sensitivity level approximately 100× higher than that of conventional protein testing. The researchers achieved rapid measurement of trace amounts on the order of tens of attograms (ag) (ag = 10−18 g; one quintillionth of a gram). They measured target protein trace amounts as small as one twenty-quadrillionth of a gram after only three minutes of irradiation.
The researchers applied the principle of light-induced acceleration to several different types of membrane proteins. In experiments, the optical technique demonstrated ultrafast, specific detection of target proteins with a smaller sample volume and higher sensitivity than conventional techniques. For example, in one type of membrane protein, the researchers detected 47 to 750 ag of target proteins, without any pretreatment, from a 300-nL sample after just three minutes of laser irradiation.
A progressive collaborative study on cancer marker measurement using patient-derived samples is underway as part of the Future Society Creation Project of the Japan Science and Technology Agency. An initial validation of the light-induced acceleration technique in clinical practice is planned, with the aim of developing a basic system within a few years, the researchers said.
Since antigen-antibody reaction is a common biochemical reaction, the technology has the potential to be used not only in the medical field but also in various industrial fields, such as testing for allergens in food and drink, detecting biological substances in the environment, and testing for intermediate products in the pharmaceutical process.
Bio Photonics Research Award
Visit: biophotonicsresearch.com
Nominate Now: https://biophotonicsresearch.com/award-nomination/?ecategory=Awards&rcategory=Awardee
#MeatAnalysis #FluorescenceTech #FoodQuality #FoodSafety #SpectroscopyInFood #MeatAuthentication #RapidDetection #FoodScience #MeatFreshness #MolecularDetection #FoodIndustryInnovation #NonDestructiveTesting #FoodMonitoring #SpectroscopyApplications #QualityControl #AdvancedSpectroscopy #MeatSpoilageDetection #FoodIntegrity #SmartFoodTesting #RealTimeAnalysis #FoodAuthenticity #FoodSafetyInnovation #SpectroscopyResearch #NextGenFoodSafety #InnovativeFoodScience,
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