Patent Application: US-42607606-A

Abstract:
the present invention provides an electrochemical biosensor test strip used for quantitative determination of an analyte in a liquid sample . methods of fabricating the test strip and reagent formulas are also provided . improvements in electrochemical test strips are required to detect the presence of a compound in a liquid mixture using a smaller sample size with increased accuracy . the present invention was developed to be user - friendly , decrease sample requirements and decrease analyzing time while increasing the reproducibility and accuracy of the electrochemical test strip .

Description:
the preferred embodiment of the present invention is shown in fig1 - 3 . fig1 illustrates an exploded perspective view of an electrochemical biosensor test strip 100 in accordance with the present invention . the electrochemical biosensor test strip 100 is a 3 - layer flat elongated construction . starting from the lowest layer , the electrochemical biosensor test strip 100 comprises a first insulating base plate 110 , then a second insulating base plate 210 , and finally a cover 310 . these layers 110 , 210 , and 310 can be made from the materials with electrically insulating property . examples of a preferred material are polyvinyl chloride , polycarbonate , polyethylene terephthalate , polyester , polystyrene , polybutadiene , polyurethane and polyimide . the first insulating base plate 110 preferably takes a rectangular form , which may be defined longitudinally as having two ends extending from a liquid sampling end 111 to an electrical contact end 112 . a pair of conductive tracks , 121 and 122 , is screen printed on the first insulating base plate 110 . near the liquid sampling end 111 are the working electrode 123 and reference electrode 124 . near the electrical contact end 112 are a plurality of electrical contacts 125 , 126 , 127 and / or 128 . the conductive tracks 121 , 122 are separated apart from each other . of the electrical contacts , at least one electrical contact ( 127 and / or 128 ) electrically connected to the electrical contact 126 is a turn - on / type - recognition electrical contact . the turn - on electrical contact is used to wake up a meter ( an apparatus to read the strip information ; not shown ) automatically when the test strip is inserted into the meter . this is used for the purpose of saving power when the battery powered meter is not in use . the type - recognition electrical contact is used to recognize the type of the strip ( e . g ., glucose strip , uric acid strip or cholesterol strip ). in other words , it is used to determine if the type of electrochemical biosensor test strip used is corresponding to the analyte to be measured , which can prevent the misusing of strips when a multi - functional meter is in user &# 39 ; s hand . suitable for use in these conductive tracks are materials with electrically conductive property . examples of a preferred material are copper , silver , gold , platinum , titanium , palladium , and carbon . the second insulating base plate 210 which overlaid on top of the first insulating base plate 110 is preferably rectangular in shape . a u - shaped cutout 211 is formed in the front end 212 of the second insulating base plate 210 . the second insulating base plate 210 has the same width as the first insulating base plate 110 . the length of the second insulating base plate 210 is shorter than that of the first insulating base plate 110 , which in turn causes an exposure of the electrical contacts 125 , 126 , 127 and / or 128 when the front end 212 is in alignment with the liquid sampling end 111 . the exposure of the electrical contacts 125 , 126 , 127 and / or 128 is used to contact the corresponding electrical contacts in the meter . the u - shaped cutout 211 of the second insulating base plate 210 not only defines the boundaries of the reaction area 113 , but it also exposes a portion of the working electrode 123 and reference electrode 124 of the test strip 100 . the preferred width and length of the u - shaped cutout 211 range from 1 mm to 4 mm and from 2 mm to 6 mm , respectively . overlaying the reaction area 113 , the exposed portion of the working electrode 123 , and reference electrode 124 , is a reagent film used to react with an analyte in a liquid sample . the reagent film is formed by drying a mixture of reagents containing at least one biological active substance ( e . g ., enzyme ) and other components such as mediator , surfactant , and at least one binder . enzymes suitable for use are ones that are responsible for the chemical reaction of the analyte . examples of a preferred enzyme are glucose oxidase or glucose dehydrogenase for measuring glucose . for measuring uric acid , the enzyme is preferably uricase . for measuring cholesterol , the enzymes are preferably cholesterol oxidase and cholesterol esterase . suitable mediator for use is the material which is capable of undergoing reversible , oxidation - reduction reaction and electron transferring . examples are potassium ferricyanide , tetrathioflilvalene , phenazine ethosulfate , and hexacyanoferrate , methylene blue , benzoquinone , and phenyidiamines , 3 , 3 ′, 5 , 5 ′- tetramethylbenzidine . surfactant suitable for use is preferably selected , either individually or in combination , from the group including triton x - 100 , cholic acid , polyethylene glycol , t - octylphenoxypolyethoxyethanol , sodium lauryl sulfate , polyoxyethylenesorbitan monolaurate ( tween 20 , tween 40 , tween 60 , tween 80 ). binders suitable for use are preferably selected , either individually or in combination , from the group including polyvinyl alcohol , polyvinyl pyrrolidone , polyethylene glycol , gelatin and methyl cellulose . the cover 310 is preferably rectangular in shape . the preferred width and length for the cover 310 is the same as the second insulating base plate 210 . when the cover 310 is placed on top of the second insulating base plate 210 , a reaction chamber is formed . the size of the reaction chamber is defined by the reaction area 113 and the thickness of the second insulating base plate 210 , which in turn defines the volume of liquid sample required per test strip . the reaction chamber of the present invention requires a lower volume of liquid sample which is about 2 . 5 microliters . the action of capillary force for introducing the liquid sample into the reaction chamber is also defined by the thickness of the second insulating base plate . acceptable thicknesses of the second insulating base plate 210 is in the range of 0 . 1 mm to 0 . 5 mm . a semicircle - shaped cutout 311 formed in the front end 312 of the cover 310 is used to accommodate a drop of user &# 39 ; s body fluid ( e . g ., finger blood ) when the front end 312 is in alignment with the front end 212 of the second insulating base plate 210 and the liquid sampling end 111 of the first insulating base plate 110 . in other words , the liquid sample is introduced into the reaction chamber from the small semicircle - shaped cutout 311 on the front end 312 of the cover 310 . the cover 310 has an open slot 315 within a picture of reverse triangle 314 . the picture of reverse triangle 314 is used as a direction instruction for inserting the test strip 100 . the open slot 315 may be used as an air vent opening to reduce the air bubbles trapped in the reaction chamber during the introduction of liquid sample . on the other hand , the open slot 315 may also be used as an indicator to insure that the reaction chamber is filled with liquid sample if the open slot 315 displays the color of the liquid sample . according to the present invention , a method for fabricating such a electrochemical biosensor test strip is provided comprising the steps of : step 1 : screen printing conductive tracks ( working electrode 123 and reference electrode 124 with a plurality of electrical contacts 125 , 126 , 127 and / or 128 ) on the first insulating base plate 110 . the conductive materials for printing such as copper , silver , gold , platinum , titanium , palladium , and carbon may be selected either individually or in combination . step 2 : after the printed conductive material is dried , a second insulating base plate 210 with a u - shaped cutout 211 in the front end 212 is placed on top of the first insulating base plate 110 . the u - shaped cutout 211 of the second insulating base plate 210 defines the boundaries of the reaction area 113 , and exposes a portion of the working electrode 123 and reference electrodes 124 of the test strip 100 . step 3 : overlaying the reaction area 113 is a reagent film which contains at least one biological active substance ( e . g ., enzyme ) and other components such as mediator , surfactant , and at least one binder . examples of the formula of the reagent film are described as follows : phosphate buffer ( ph = 6 . 5 ) 87 . 0 % potassium ferricyanide 10 . 0 % glucose oxidase 1 . 2 % methyl cellulose 1 . 0 % triton x - 100 0 . 6 % bovine serum albumin 0 . 2 % phosphate buffer ( ph = 6 . 5 ) 87 . 0 % potassium ferricyanide 10 . 3 % glucose oxidase 1 . 4 % peg 0 . 6 % tween 20 0 . 4 % bovine serum albumin 0 . 3 % phosphate buffer ( ph = 6 . 5 ) 87 . 0 % potassium ferricyanide 10 . 0 % glucose oxidase 1 . 2 % carboxymethyl cellulose 1 . 0 % triton x - 100 0 . 6 % bovine serum albumin 0 . 2 % phosphate buffer ( ph = 7 . 0 ) 92 . 4 % potassium ferricyanide 6 . 0 % uricase 0 . 4 % methyl cellulose 0 . 6 % triton x - 100 0 . 5 % bovine serum albumin 0 . 1 % phosphate buffer ( ph = 7 . 0 ) 84 . 8 % potassium ferricyanide 10 . 0 % cholesterol esterase 1 . 0 % cholesterol oxidase 2 . 0 % methyl cellulose 1 . 0 % triton x - 100 1 . 0 % bovine serum albumin 0 . 2 % step 4 : placing a cover 310 on top of the second insulating base plate 210 and aligning two front ends 312 , 212 — an electrochemical biosensor test strip is ready for use . when a liquid sample is introduced to a single electrochemical biosensor test strip of the present invention , the liquid sample enters the reaction chamber from the small semicircle - shaped cutout 311 on the front end 312 of the cover 310 and stops at bottom end of the u - shaped cutout 211 of the second insulating base plate 210 . when the color of the liquid sample is shown on the open slot 315 , the reaction chamber is filled with liquid sample . the volume of the liquid sample required for the electrochemical biosensor test strip is about 2 . 5 microliters . the following example illustrates the correlation of glucose concentration readings determined by the test strips of the present invention and by a traditional ysi glucose analyzer . to make clinically relevant blood glucose concentrations , a venous blood sample was collected , separated into 6 tubes , and each tube was spiked with different glucose concentrations ( 43 , 91 , 176 , 234 , 354 , and 449 mg / dl ). these blood glucose concentrations were determined by a ysi glucose analyzer . using the ebsensor glucose meter ( not shown ), the coefficients of variation ( cvs ) calculated from the 30 measurements of each glucose concentration were found to be 5 . 87 %, 5 . 46 %, 3 . 99 %, 4 . 90 %, 4 . 64 %, and 5 . 61 %, respectively . the glucose concentration readings using electrochemical biosensor test strips of the present invention plotted against the concentration values determined by the ysi analyzer are illustrated in fig4 . a regression coefficient of 0 . 99 indicated a linear relationship of glucose concentration readings using the test strips of the present invention versus glucose concentration readings determined by the ysi analyzer . all references are listed herein for the convenience of the reader . each is incorporated by reference in its entirety . 1 . u . s . pat . no . 5 , 120 , 420 biosensor and a process for preparation thereof , ( nankai et al . 1992 ). 2 . u . s . pat . no . 5 , 288 , 636 enzyme electrode system , ( pollmann et al ., 1994 ) 3 . u . s . pat . no . 5 , 437 , 999 electrochemical sensor , ( diebold et al . 1995 ) 4 . u . s . pat . no . 5 , 727 , 548 strip electrode with screen printing , ( hill et al . 1998 ) 5 . u . s . pat . no . 5 , 762 , 770 electrochemical biosensor test strip , ( pritchard et al . 1998 ) 6 . alto et al . 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