Lateral flow test strip assay for osteoporosis

Abstract

A lateral flow test strip (LFTS) platform measures osteocalcin (OC) in saliva to identify early indications of bone loss and minimize bone fracture risk associated with osteoporosis. The OC assay embodiments are based on the experimentally identified optimal markers which exhibit selectivity with very low false positives, and sensitivity relevant to clinical requirements. A prospective clinical study sampling of 20 patients demonstrated excellent correlation of OC in saliva with bone mineral density (BMD). Salivary OC and Dpd levels were validated with a standard commercial ELISA kit against serum (OC) and urine (Dpd). Multiplexed LFTS are used to increase specificity of an assay.

What is claimed is: 
     
         1 . A lateral flow test strip for screening for osteoporosis comprising:
 a) a sample pad for accepting a subject sample bodily fluid;   b) a migration membrane;   c) a plastic backing;   d) a conjugate pad deposited with a first fluorescently labeled detector antibody;   e) a first capture line comprising a first capture antibody specific to osteocalcin wherein a first antibody pair comprising the first capture antibody and the first fluorescently labeled detector antibody are chosen from the group consisting of:
 i. 940/939, 
 ii. 11F8/939, 
 iii. 2H9/939, 
 iv. 940/H10, 
 v. 2H9/H10, 
 vi. 940/3G8, 
 vii. 11F8/3G8. 
   
     
     
         2 . The lateral flow test strip of  claim 1  also comprising a second capture line comprising a second capture antibody reactive specific to:
 a) deoxypyridinoline, 
 b) urinary calcium, 
 c) tartare resistant acid phosphtase, 
 d) bone sialoprotein, 
 e) pryidinoline, 
 f) N-telopeptide, 
 g) C-telopeptide, 
 h) C-terminal telopeptide of type I collagen 
 i) bone-specific alkaline phosphatase, 
 j) pro-collagen I extension peptides, 
 k) amino terminal. 
 
     
     
         3 . The lateral flow test strip of  claim 1  wherein the conjugate pad deposited with the first fluorescently labeled detector antibody is conjugated to fluorescent microparticles being paired with the first capture antibody being immobilized on the migration membrane. 
     
     
         4 . The lateral flow test strip of  claim 1  wherein the bodily fluid is whole saliva. 
     
     
         5 . The lateral flow test strip of  claim 1  wherein the bodily fluid is urine. 
     
     
         6 . The lateral flow test strip of  claim 1  also comprising a second capture line comprising a second capture antibody specific to osteocalcin wherein a second antibody pair comprising the second capture antibody and a second fluorescently labeled detector antibody are chosen from the group consisting of:
 a) 940/939, 
 b) 11F8/939, 
 c) 2H9/939, 
 d) 940/H10, 
 e) 2H9/H10, 
 f) 940/3G8, 
 g) 11F8/3G8. 
 
     
     
         7 . The lateral flow test strip of  claim 6  wherein the first capture antibody and the second capture antibody are the same. 
     
     
         8 . A system for immunochromatographic bone loss marker assay comprising:
 a) a lateral flow test strip comprising:
 i) a sample pad for accepting a subject sample bodily fluid; 
 ii) a migration membrane; 
 iii) a plastic backing; 
 iv) a conjugate pad deposited with a first fluorescently labeled detector antibody; 
 v) a first capture line comprising a first capture antibody specific to osteocalcin wherein the first capture antibody and the first fluorescently labeled detector antibody are chosen from the group consisting of:
 (1) 940/939, 
 (2) 11F8/939, 
 (3) 2H9/939, 
 (4) 940/H10, 
 (5) 2H9/H10, 
 (6) 940/3G8, 
 (7) 11F8/3G8. 
 
   b) a calibrated florescence test strip reader.   
     
     
         9 . The system of  claim 8  wherein the conjugate pad deposited with the first fluorescently labeled detector antibody is conjugated to fluorescent microparticles being paired with the first capture antibody being immobilized on the migration membrane. 
     
     
         10 . The system of  claim 8  wherein the lateral flow test strip also comprises a second capture line comprising a second antibody capture antibody reactive specific to:
 a) deoxypyridinoline, 
 b) urinary calcium, 
 c) tartare resistant acid phosphtase, 
 d) bone sialoprotein, 
 e) pryidinoline, 
 f) N-telopeptide, 
 g) C-telopeptide, 
 h) C-terminal telopeptide of type I collagen 
 i) bone-specific alkaline phosphatase, 
 j) pro-collagen I extension peptides, 
 k) amino terminal. 
 
     
     
         11 . The system of  claim 8  wherein the bodily fluid is whole saliva. 
     
     
         12 . The system of  claim 8  wherein the bodily fluid is urine. 
     
     
         13 . The system of  claim 8  wherein the immunochromatographic assay also comprises a diagnostically correlated evaluation for reader output measurements of reagent lines following the application of a serum specimen to the lateral flow test strip. 
     
     
         14 . A multi-plexed lateral flow test strip for screening for osteoporosis comprising:
 a) a sample pad for accepting a subject sample bodily fluid;   b) a migration membrane;   c) a plastic backing;   d) a conjugate pad deposited with a first fluorescently labeled detector antibody;   e) a first capture line comprising a first capture antibody specific to osteocalcin wherein a first antibody pair comprising the first capture antibody and the first fluorescently labeled detector antibody are chosen from the group consisting of:   f) a second capture line comprising a second capture antibody specific to secondary marker for osteocalcin wherein a second antibody pair comprise a second capture antibody and a second fluorescently labeled detector antibody. are chosen from the group consisting of
 i. osteocalcin, 
 ii. deoxypyridinoline, 
 iii. urinary calcium, 
 iv. tartare resistant acid phosphtase, 
 v. bone sialoprotein, 
 vi. pryidinoline, 
 vii. N-telopeptide, 
 viii. C-telopeptide, 
 ix. C-terminal telopeptide of type I collagen 
 x. bone-specific alkaline phosphatase, 
 xi. pro-collagen I extension peptides, 
 xii. amino terminal.