Evaluation of human thymus and activation-regulated chemokine concentrations in blood using a new sandwich ELISA based on monoclonal antibodies.
BACKGROUND
CC chemokine TARC (thymus and activation-regulated chemokine), a potent chemoattractant for Th2 lymphocytes, is thought to play important roles in inflammatory diseases. We developed a new sensitive enzyme-linked immunoassay (ELISA) for human TARC (hTARC) to accurately measure and evaluate its concentrations in blood.


METHODS
An ELISA was developed using two established monoclonal antibodies against hTARC. Using this assay, we observed changes of hTARC concentrations in serum and plasma obtained from individual subjects. Improvements to the assay were made to allow use for the clinical evaluation of samples from atopic dermatitis (AD).


RESULTS
The lower detection limit of the ELISA was 1.4 pg/ml for a 25 microl sample volume. Other assay characteristics were enough to satisfactorily measure hTARC in biological fluids. This ELISA revealed that changes in serum and plasma concentrations were related to sample handling before separation from blood. With appropriate sample preparation, significant increases of hTARC were observed in patients with AD in comparison with normal subjects.


CONCLUSIONS
Appropriate sample preparation is important for clinical studies on hTARC. Accurate measurement using our ELISA method offers a suitable clinical index for evaluating the severity of allergic diseases of Th2-dominant disorders, such as AD.
Morita
Atsushi
A
Diagnostics Department, Shionogi & Co., Ltd., 2-5-1 Mishima, Settsu-shi, Osaka 566-0022, Japan. atsushi.morita@shionogi.co.jp
Kikuoka
Shino
S
Horikawa
Tatsuya
T
Bito
Toshinori
T
Yamada
Hirokazu
H
Kanda
Megumi
M
Sasakura
Kazuyuki
K
Tamaki
Mikio
M
Hirai
Koichi
K
Suzuki
Ryuji
R
Sugita
Kenji
K
D000911:Antibodies, Monoclonal / Q000276:immunology*; D002138:Calibration; D054414:Chemokine CCL17; D019742:Chemokines, CC / Q000097:blood* / Q000276:immunology*; D003876:Dermatitis, Atopic / Q000097:blood; D004492:Edetic Acid; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D012680:Sensitivity and Specificity; D013997:Time Factors
D000911:Antibodies, Monoclonal; C106230:CCL17 protein, human; D054414:Chemokine CCL17; D019742:Chemokines, CC; D004492:Edetic Acid