Two-step sandwich enzyme immunoassay using monoclonal antibodies for detection of soluble and membrane-associated human membrane Type 1-matrix metalloproteinase

Takanori Aoki, Kayoko Yonezawa, Eiko Ohuchi, Noboru Fujimoto, Kazushi Iwata, Taketoshi Shimada, Takayuki Shiomi, Yasunori Okada, Motoharu Seiki

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

A two-step sandwich enzyme immunoassay (EIA) system for the detection of human membrane Type 1-matrix metalloproteinase (MT1-MMP) was established by using two monoclonal antibodies against recombinant MT1-MMP. MT1-MMP in which samples were reacted with solid-phase antibody and then detected with peroxidase-labeled second antibody. At least 1.25 ng/mL was detected by the EIA system, and linearity was obtained between 1.25 and 160 ng/mL. This EIA system is specific for MT1-MMP and did not show cross-reactivity against several other MMP's examined. Shedding of soluble MT1-MMP into the medium by some cancer cell lines was also detected by this system. However, soluble MT1-MMP in serum from normal and cancer patients was under the detection limit. Membrane-associated MT1-MMP of cancer cell lines was also detected after solubilization of the membranes with extraction buffer containing detergent. Additionally, MT1-MMP in clinical samples was examined. Elevated levels of MT1-MMP were detected in homogenate of cancer tissue compared with the levels for normal tissue and the level of MT1-MMP in tumors correlated with the rate of metastasis to the regional lymph nodes. Thus, we demonstrated that this EIA system is the first to measure MT1-MMP in clinical specimens, thus suggesting its useful for diagnosis of cancer or prediction of malignancy.

Original languageEnglish
Pages (from-to)49-68
Number of pages20
JournalJournal of Immunoassay and Immunochemistry
Volume23
Issue number1
DOIs
Publication statusPublished - 2002

Fingerprint

Matrix Metalloproteinase 14
Immunoenzyme Techniques
Monoclonal Antibodies
Membranes
Enzymes
Neoplasms
Membrane-Associated Matrix Metalloproteinases
human MMP14 protein
Cells
Tissue
Cell Line
Antibodies
Matrix Metalloproteinases
Detergents
Peroxidase
Limit of Detection
Tumors
Buffers
Lymph Nodes

ASJC Scopus subject areas

  • Immunology
  • Pharmacology

Cite this

Two-step sandwich enzyme immunoassay using monoclonal antibodies for detection of soluble and membrane-associated human membrane Type 1-matrix metalloproteinase. / Aoki, Takanori; Yonezawa, Kayoko; Ohuchi, Eiko; Fujimoto, Noboru; Iwata, Kazushi; Shimada, Taketoshi; Shiomi, Takayuki; Okada, Yasunori; Seiki, Motoharu.

In: Journal of Immunoassay and Immunochemistry, Vol. 23, No. 1, 2002, p. 49-68.

Research output: Contribution to journalArticle

Aoki, Takanori ; Yonezawa, Kayoko ; Ohuchi, Eiko ; Fujimoto, Noboru ; Iwata, Kazushi ; Shimada, Taketoshi ; Shiomi, Takayuki ; Okada, Yasunori ; Seiki, Motoharu. / Two-step sandwich enzyme immunoassay using monoclonal antibodies for detection of soluble and membrane-associated human membrane Type 1-matrix metalloproteinase. In: Journal of Immunoassay and Immunochemistry. 2002 ; Vol. 23, No. 1. pp. 49-68.
@article{5041fd1a7132471eb3e3e4c5869ce517,
title = "Two-step sandwich enzyme immunoassay using monoclonal antibodies for detection of soluble and membrane-associated human membrane Type 1-matrix metalloproteinase",
abstract = "A two-step sandwich enzyme immunoassay (EIA) system for the detection of human membrane Type 1-matrix metalloproteinase (MT1-MMP) was established by using two monoclonal antibodies against recombinant MT1-MMP. MT1-MMP in which samples were reacted with solid-phase antibody and then detected with peroxidase-labeled second antibody. At least 1.25 ng/mL was detected by the EIA system, and linearity was obtained between 1.25 and 160 ng/mL. This EIA system is specific for MT1-MMP and did not show cross-reactivity against several other MMP's examined. Shedding of soluble MT1-MMP into the medium by some cancer cell lines was also detected by this system. However, soluble MT1-MMP in serum from normal and cancer patients was under the detection limit. Membrane-associated MT1-MMP of cancer cell lines was also detected after solubilization of the membranes with extraction buffer containing detergent. Additionally, MT1-MMP in clinical samples was examined. Elevated levels of MT1-MMP were detected in homogenate of cancer tissue compared with the levels for normal tissue and the level of MT1-MMP in tumors correlated with the rate of metastasis to the regional lymph nodes. Thus, we demonstrated that this EIA system is the first to measure MT1-MMP in clinical specimens, thus suggesting its useful for diagnosis of cancer or prediction of malignancy.",
author = "Takanori Aoki and Kayoko Yonezawa and Eiko Ohuchi and Noboru Fujimoto and Kazushi Iwata and Taketoshi Shimada and Takayuki Shiomi and Yasunori Okada and Motoharu Seiki",
year = "2002",
doi = "10.1081/IAS-120002274",
language = "English",
volume = "23",
pages = "49--68",
journal = "Journal of Immunoassay and Immunochemistry",
issn = "1532-1819",
publisher = "Taylor and Francis Ltd.",
number = "1",

}

TY - JOUR

T1 - Two-step sandwich enzyme immunoassay using monoclonal antibodies for detection of soluble and membrane-associated human membrane Type 1-matrix metalloproteinase

AU - Aoki, Takanori

AU - Yonezawa, Kayoko

AU - Ohuchi, Eiko

AU - Fujimoto, Noboru

AU - Iwata, Kazushi

AU - Shimada, Taketoshi

AU - Shiomi, Takayuki

AU - Okada, Yasunori

AU - Seiki, Motoharu

PY - 2002

Y1 - 2002

N2 - A two-step sandwich enzyme immunoassay (EIA) system for the detection of human membrane Type 1-matrix metalloproteinase (MT1-MMP) was established by using two monoclonal antibodies against recombinant MT1-MMP. MT1-MMP in which samples were reacted with solid-phase antibody and then detected with peroxidase-labeled second antibody. At least 1.25 ng/mL was detected by the EIA system, and linearity was obtained between 1.25 and 160 ng/mL. This EIA system is specific for MT1-MMP and did not show cross-reactivity against several other MMP's examined. Shedding of soluble MT1-MMP into the medium by some cancer cell lines was also detected by this system. However, soluble MT1-MMP in serum from normal and cancer patients was under the detection limit. Membrane-associated MT1-MMP of cancer cell lines was also detected after solubilization of the membranes with extraction buffer containing detergent. Additionally, MT1-MMP in clinical samples was examined. Elevated levels of MT1-MMP were detected in homogenate of cancer tissue compared with the levels for normal tissue and the level of MT1-MMP in tumors correlated with the rate of metastasis to the regional lymph nodes. Thus, we demonstrated that this EIA system is the first to measure MT1-MMP in clinical specimens, thus suggesting its useful for diagnosis of cancer or prediction of malignancy.

AB - A two-step sandwich enzyme immunoassay (EIA) system for the detection of human membrane Type 1-matrix metalloproteinase (MT1-MMP) was established by using two monoclonal antibodies against recombinant MT1-MMP. MT1-MMP in which samples were reacted with solid-phase antibody and then detected with peroxidase-labeled second antibody. At least 1.25 ng/mL was detected by the EIA system, and linearity was obtained between 1.25 and 160 ng/mL. This EIA system is specific for MT1-MMP and did not show cross-reactivity against several other MMP's examined. Shedding of soluble MT1-MMP into the medium by some cancer cell lines was also detected by this system. However, soluble MT1-MMP in serum from normal and cancer patients was under the detection limit. Membrane-associated MT1-MMP of cancer cell lines was also detected after solubilization of the membranes with extraction buffer containing detergent. Additionally, MT1-MMP in clinical samples was examined. Elevated levels of MT1-MMP were detected in homogenate of cancer tissue compared with the levels for normal tissue and the level of MT1-MMP in tumors correlated with the rate of metastasis to the regional lymph nodes. Thus, we demonstrated that this EIA system is the first to measure MT1-MMP in clinical specimens, thus suggesting its useful for diagnosis of cancer or prediction of malignancy.

UR - http://www.scopus.com/inward/record.url?scp=0036169341&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0036169341&partnerID=8YFLogxK

U2 - 10.1081/IAS-120002274

DO - 10.1081/IAS-120002274

M3 - Article

C2 - 11848101

AN - SCOPUS:0036169341

VL - 23

SP - 49

EP - 68

JO - Journal of Immunoassay and Immunochemistry

JF - Journal of Immunoassay and Immunochemistry

SN - 1532-1819

IS - 1

ER -