Point-of-Care Fluorescent Immunoassay for Identifying Biomarkers in Patient Biofluid Samples

Abstract

Systems and methods for low-cost point-of-care immunoassay are provided. The system comprises an emitter, two optical interference filters, a microscope slide, a photodiode detector, a circuit, and a measuring unit. The detector is placed upon the second filter, the slide, the first filter, and then the emitter. The emitter comprises non-organic light emitting diodes (LEDs) or organic light emitting diodes (OLEDs) that emits light of a first color. The slide is spotted with biofluid from a patient. Biomarkers in the biofluid is bound with immobilized fluorophores that emit light of a second color when stimulated by the light of the first color. The first and second filters band-pass the light of the first and second colors, respectively. The detector detects light of the second color. The current outputted from the detector is converted into a relatively-large output voltage by a circuit. A measuring unit measures the ramp time of the output voltage. The ramp time is to be used to determine the concentration of the fluorophores, which in turn is related to the concentration of the biomarkers.

We claim: 
     
         1 . A method for providing low-cost point-of-care quantitative diagnostic information, comprising:
 a) preparing a microscope slide spotted with biofluid from a patient, wherein fluorophores are immobilized and bound with biomarkers in the biofluid;   b) assembling a layered structure comprising
 a LED or OLED emitter, 
 a first optical interference filter, 
 the slide, 
 a second optical interference filter, and 
 a photodiode detector, wherein:
 the slide is sandwiched between the first filter and the second filter, 
 the first filter is placed over the emitter and band-passes light of a first color emitted from the emitter, 
 the fluorophores emit light of a second color when stimulated by the light of the first color, 
 the second filter band-passes the light of the second color, and 
 the detector is placed over the second filter and detects the light of the second color; 
 
   c) detect current caused by the light of the second color using the detector;   d) converting the current into a relatively-large output voltage;   d) measuring a ramp time of the output voltage;   e) determining a concentration of the fluorophores related to a concentration of the biomarkers using the ramp time; and   f) generating a report about the biomarkers using the concentration of the fluorophores.   
     
     
         2 . The method as recited in  claim 1 , wherein a 3D printed assembly is used and designed to align a center of the slide with the first filter and the second filter. 
     
     
         3 . The method as recited in  claim 1 , wherein step a) includes:
 i) silanizing the slide;   ii) spotting captured proteins onto the slide;   iii) incubating the proteins with the biofluid; and   iv) incubating the proteins further with detection antibodies associated with the biomarkers.   
     
     
         4 . The method as recited in  claim 3 , wherein step ii) is repeated one or more times to achieve a desired sensitivity. 
     
     
         5 . The method as recited in  claim 1 , wherein the ramp time is a time interval that the output voltage takes to ramp from 0 volt to a supply voltage rail. 
     
     
         6 . The method as recited in  claim 1 , wherein the ramp time is a time interval that the output voltage takes to ramp from a first voltage to a second voltage. 
     
     
         17 . The method as recited in  claim 1 , wherein the first filter is deposited in the emitter. 
     
     
         8 . The method as recited in  claim 1 , wherein the biofluid is serum. 
     
     
         9 . The method as recited in  claim 8 , wherein the biomarkers are for papillomavirus. 
     
     
         10 . The method as recited in  claim 1 , wherein the biomarkers are antibodies specific for HPV16 E7. 
     
     
         11 . The method as recited in  claim 1 , wherein the report indicates positive or negative with the biomarkers based on three standard deviations of a mean of controls. 
     
     
         12 . A method of immunoassay to be used for identifying biomarkers in a sample of patient biofluid, comprising:
 i) silanizing a microscope slide;   ii) spotting captured proteins onto the slide;   iii) incubating the proteins with the patient biofluid; and   iv) incubating the proteins further with detection antibodies associated with the biomarkers.   
     
     
         13 . The method as recited in  claim 12 , wherein step ii) is repeated one or more times to achieve desired sensitivity.