Generation and characterization of monoclonal antibodies to human type-5 tartrate-resistant acid phosphatase: development of a specific immunoassay of the isoenzyme in serum

Abstract We have characterized four monoclonal antibodies (mAbs) to the purple ("tartrate-resistant," band 5) acid phosphatase of the human osteoclast (TRAP) and used these to develop a specific serum immunoassay. All four mAbs are of high affinity (Ka = 1-5 x 10(8) L/mol) with a very fast Kassoc (0.2-2.0 x 10(5) L mol-1 s-1) and a moderate Kdissoc (1-3 x 10(-3) s). Two of the mAbs were selected to develop a time-resolved fluorescence immunoassay to measure serum concentrations of TRAP. The mean serum immunoreactive TRAP in a group of healthy premenopausal women and men was 3.7 +/- 1.8 micrograms/L (mean +/- SD) and 3.5 +/- 1.6 micrograms/L, respectively. Significantly higher concentrations of TRAP were found in postmenopausal women (6.3 +/- 2.3 micrograms/L) and in eight patients with Gaucher disease (19.3 +/- 4.7 micrograms/L). Further studies are required to investigate the value of serum TRAP as a marker of bone resorption.

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Development and Characterization of Novel Monoclonal Antibodies Against Tartrate-Resistant Acid Phosphatase 5

Hybridoma ◽

10.1089/hyb.2006.25.358 ◽

2006 ◽

Vol 25 (6) ◽

pp. 358-366 ◽

Cited By ~ 1

Author(s):

Tatsuya Ohashi ◽

Toshihide Miura ◽

Yoshihiko Igarashi ◽

Iwao Kiyokawa ◽

Yasuhito Sato ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Tartrate Resistant Acid Phosphatase

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Characterization of Monoclonal Antibodies Specific to Human Tartrate-Resistant Acid Phosphatase

Hybridoma ◽

10.1089/hyb.1997.16.175 ◽

1997 ◽

Vol 16 (2) ◽

pp. 175-182 ◽

Cited By ~ 7

Author(s):

ANTHONY J. JANCKILA ◽

ERNEST M. CARDWELL ◽

LUNG T. YAM

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Tartrate Resistant Acid Phosphatase

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Two-Site Immunoassays for Osteoclastic Tartrate-Resistant Acid Phosphatase Based on Characterization of Six Monoclonal Antibodies

Journal of Bone and Mineral Research ◽

10.1359/jbmr.1999.14.3.464 ◽

1999 ◽

Vol 14 (3) ◽

pp. 464-469 ◽

Cited By ~ 33

Author(s):

Jussi M. Halleen ◽

Matti Karp ◽

Sari Viloma ◽

Pirjo Laaksonen ◽

Jukka Hellman ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Tartrate Resistant Acid Phosphatase

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Characterization of Four Monoclonal Antibodies to Recombinant Human Tartrate-Resistant Acid Phosphatase

Hybridoma and Hybridomics ◽

10.1089/153685902760173917 ◽

2002 ◽

Vol 21 (3) ◽

pp. 191-195 ◽

Cited By ~ 3

Author(s):

Takashi Miyazaki ◽

Toshiyuki Matsunaga ◽

Shuichi Miyazaki ◽

Shigeru Hokari ◽

Tsugikazu Komoda

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Tartrate Resistant Acid Phosphatase

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Biochemical characterization of the tartrate-resistant acid phosphatase of human spleen with leukemic reticuloendotheliosis as a pyrophosphatase.

Clinical Chemistry ◽

10.1093/clinchem/23.1.89 ◽

1977 ◽

Vol 23 (1) ◽

pp. 89-94 ◽

Cited By ~ 29

Author(s):

K W Lam ◽

L T Yam

Keyword(s):

Acid Phosphatase ◽

Biochemical Characterization ◽

Normal Animal ◽

Tartrate Resistant Acid Phosphatase ◽

Animal Tissues ◽

Human Spleen ◽

Optimal Activity ◽

Catalytic Function ◽

Hydrolysis Of

Abstract A tartrate-resistant acid phosphatase was isolated from a human leukemic spleen by freeze-thawing in saline and purified by repeated chromatography on carboxymethyl-cellulose. The purified enzyme has a molecular weight of 64 000. It catalyzes the hydrolysis of inorganic and organic pyrophosphate as well as the phenolic ester of monoorthophosphate, with optimal activity between pH 5 and 6. However, there is no activity toward mono-orthophosphate esters of aliphatic alcohols. The present data have identified its catalytic function as a pyrophosphatase. However, it has properties different from the pyrophosphatase previously observed in normal animal tissues.

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Characterization of mineral deposits formed in cultures of a hamster tartrate-resistant acid phosphatase (TRAP) and alkaline phosphatase (ALP) double-positive cell line (CCP)

Cell and Tissue Research ◽

10.1007/s00441-002-0587-y ◽

2002 ◽

Vol 309 (2) ◽

pp. 269-279 ◽

Cited By ~ 3

Author(s):

Hisako Sakiyama ◽

Takashi Nonaka ◽

Yoshinori Kuboki ◽

Yoshio Hirabayasi ◽

Kazuko Yoshida ◽

...

Keyword(s):

Alkaline Phosphatase ◽

Acid Phosphatase ◽

Cell Line ◽

Mineral Deposits ◽

Tartrate Resistant Acid Phosphatase ◽

Double Positive ◽

Double Positive Cell

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A Novel Sandwich ELISA for Tartrate-Resistant Acid Phosphatase 5a and 5b Protein Reveals that Both Isoforms are Secreted by Differentiating Osteoclasts and Correlate to the Type I Collagen Degradation Marker CTX-I In Vivo and In Vitro

Calcified Tissue International ◽

10.1007/s00223-019-00618-w ◽

2019 ◽

Vol 106 (2) ◽

pp. 194-207 ◽

Cited By ~ 1

Author(s):

Laia Mira-Pascual ◽

Christina Patlaka ◽

Suchita Desai ◽

Staffan Paulie ◽

Tuomas Näreoja ◽

...

Keyword(s):

Acid Phosphatase ◽

Bone Resorption ◽

Sandwich Elisa ◽

Osteoclast Differentiation ◽

Collagen Degradation ◽

Considerable Proportion ◽

Tartrate Resistant Acid Phosphatase ◽

Osteoclast Number ◽

Serum Trap ◽

Trap 5B

Abstract Tartrate-resistant acid phosphatase type 5 (TRAP) exists as two isoforms, 5a and 5b. 5b is a marker of osteoclast number and 5a of chronic inflammation; however, its association with bone resorption is unknown. In this study, a double-TRAP 5a/5b sandwich ELISA measuring 5a and 5b protein in the same sample was developed. TRAP 5a and 5b protein levels were evaluated as osteoclast differentiation/activity markers in serum and in culture, and their correlation to the resorption marker CTX-I was examined. Serum TRAP 5a and 5b concentrations in healthy men were 4.4 ± 0.6 ng/ml and 1.3 ± 0.2 ng/ml, respectively, and they correlated moderately to each other suggesting that their secretion is coupled under healthy conditions. A correlation was also observed between serum TRAP 5a and 5b with CTX-I, suggesting that both TRAP isoforms associate with osteoclast number. During osteoclast differentiation on plastic/bone, predominantly 5b increased in media/lysate from M-CSF/RANKL-stimulated CD14+ PBMCs. However, substantial levels of 5a were detected at later stages suggesting that both isoforms are secreted from differentiating OCs. More TRAP 5b was released on bone indicating a connection to osteoclast resorptive activity, and a peak in TRAP 5b/5a-ratio coincided with rapid CTX-I release. At the end of the culture period of M-CSF + RANKL-stimulated CD14+ PBMCs, there was a correlation between the secretion of TRAP 5a and 5b proteins with CTX-I. The correlation of not only 5b but also 5a with collagen degradation, both in serum and osteoclast cultures indicates that a considerable proportion of the TRAP 5a originates from osteoclasts and may reflect a hitherto undisclosed regulatory mechanism during bone resorption and bone remodeling.

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Clinical Significance of Immunoassays for Type-5 Tartrate-resistant Acid Phosphatase

Clinical Chemistry ◽

10.1093/clinchem/45.12.2150 ◽

1999 ◽

Vol 45 (12) ◽

pp. 2150-2157 ◽

Cited By ~ 38

Author(s):

Yuri R Nakasato ◽

Anthony J Janckila ◽

Jussi M Halleen ◽

H Kalervo Vaananen ◽

Stephanie P Walton ◽

...

Keyword(s):

Acid Phosphatase ◽

Rheumatic Disease ◽

Healthy Subjects ◽

Specific Activity ◽

Biological Significance ◽

Tartrate Resistant Acid Phosphatase ◽

Endstage Renal Disease ◽

Trap Activity ◽

Specificity And Sensitivity ◽

The Mean

Abstract Background: Tartrate-resistant acid phosphatase (TRAP; EC 3.1.3.2) is a product of osteoclasts and a biochemical marker of bone resorption rate. However, erythrocytes and platelets contribute to total TRAP activity in serum, reducing the specificity of direct biochemical assays in serum. Osteoclast TRAP is also known as type-5 TRAP and is antigenically unique. Immunoassays are sought to improve the specificity and sensitivity of TRAP as a bone marker. Methods: We developed two colorimetric microplate assays for type-5 TRAP: an enzyme capture immunoassay to measure antibody-bound enzymatic activity, and a two-site immunoassay to measure bound enzyme protein. Both use the same monoclonal antibody (14G6) to capture type-5 TRAP, which permits determination of specific activity of serum TRAP in health and disease. Results: Both TRAP assays were linear from one-tenth to fivefold the mean value in 18 healthy subjects. In these subjects, the mean (SD) TRAP activity was 3.2 (0.54) U/L for the enzyme capture assay and 37 (13) μg/L for the two-site assay. Mean TRAP activity was not significantly increased in 64 patients with endstage renal disease requiring hemodialysis (HD) or 99 unselected patients with rheumatic diseases. By contrast, TRAP protein was increased in both the HD and rheumatic disease groups. The specific activity of TRAP in the 17 of 64 HD sera that had increased TRAP activity (0.088 U/μg) was similar to that in healthy subjects (0.091 U/μg). By contrast, the specific activity of TRAP in the 31 of 99 rheumatic sera with increased TRAP protein (0.035 U/μg) was significantly decreased. Conclusions: Wide sample distributions for TRAP activity in HD patients and TRAP protein in rheumatic disease patients suggest the presence of subpopulations of HD patients with increased TRAP activity and of rheumatic patients with increased TRAP protein. Each assay for TRAP activity and protein may have its own biological significance and clinical applications in specific groups of patients.

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