Quantification of cystatin C by time-resolved fluorometry-based immunoassays
Ristiniemi, Noora ; Qin, Qiu-Ping ; Lindström, Veronica LU
; Grubb, Anders
LU
and Pettersson, Kim
(2012)
In Journal of Immunological Methods
378(1-2).
p.56-61
- Abstract
- Plasma cystatin C is increasingly used as a marker of glomerular filtration rate. Most assays for cystatin C are based on turbidimetric or nephelometric detection and studies of other rapid methods are limited. This study aimed to develop and compare differently configured immunoassays for quantification of plasma cystatin C, using recombinant cystatin C and two cystatin C-specific antibodies. Method 1 was a two-step sandwich assay with polyclonal antibody as capture and europium chelate-labeled monoclonal antibody as tracer. Method 2 was a one-step heterogeneous competitive assay using immobilized polyclonal antibody and europium-labeled cystatin C. Method 3 was a one-step homogeneous competitive assay with europium-labeled polyclonal... (More)
- Plasma cystatin C is increasingly used as a marker of glomerular filtration rate. Most assays for cystatin C are based on turbidimetric or nephelometric detection and studies of other rapid methods are limited. This study aimed to develop and compare differently configured immunoassays for quantification of plasma cystatin C, using recombinant cystatin C and two cystatin C-specific antibodies. Method 1 was a two-step sandwich assay with polyclonal antibody as capture and europium chelate-labeled monoclonal antibody as tracer. Method 2 was a one-step heterogeneous competitive assay using immobilized polyclonal antibody and europium-labeled cystatin C. Method 3 was a one-step homogeneous competitive assay with europium-labeled polyclonal antibody as donor and cyanine 5-labeled cystatin C as acceptor. All three assays were evaluated with plasma samples and their performance was compared to a conventional particle-enhanced turbidimetric immunoassay (PETIA). Method 3 was the easiest to perform, with incubation at ambient temperature for 10 min and 20 mu L of sample, while methods 1 and 2 had washing steps, took 40 min and 15 min at 37 degrees C, respectively, but used only 10 mu L of 100- or 10-fold diluted sample, respectively. The working ranges for methods 1, 2 and 3 were 0.0005-0.2, 0.05-1.0 and 0.25-20 mg/L, respectively. Kinetics for method 3 was the fastest with >95% binding completion and for method 2 the slowest with 60% binding completion. All three methods showed good correlation to PETIA, but produced higher cystatin C levels than PETIA. Methods 1 and 3 offered the most favorable performance characteristics and especially method 3 enabled rapid and simple measurement of circulating cystatin C. (C) 2012 Elsevier B.V. All rights reserved. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/2809703
- author
- Ristiniemi, Noora
; Qin, Qiu-Ping
; Lindström, Veronica
LU
; Grubb, Anders
LU
and Pettersson, Kim
- organization
- publishing date
- 2012
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Cystatin C, Immunoassay, Heterogeneous assay, Homogeneous assay, Time-resolved fluorescence
- in
- Journal of Immunological Methods
- volume
- 378
- issue
- 1-2
- pages
- 56 - 61
- publisher
- Elsevier
- external identifiers
-
- wos:000304285900007
- scopus:84859425466
- pmid:22349125
- ISSN
- 1872-7905
- DOI
- 10.1016/j.jim.2012.02.004
- language
- English
- LU publication?
- yes
- id
- 4155d9c5-cea1-4663-aaa6-96f6fb8f9a1a (old id 2809703)
- date added to LUP
- 2016-04-01 14:46:11
- date last changed
- 2023-01-04 06:34:30
@article{4155d9c5-cea1-4663-aaa6-96f6fb8f9a1a,
abstract = {{Plasma cystatin C is increasingly used as a marker of glomerular filtration rate. Most assays for cystatin C are based on turbidimetric or nephelometric detection and studies of other rapid methods are limited. This study aimed to develop and compare differently configured immunoassays for quantification of plasma cystatin C, using recombinant cystatin C and two cystatin C-specific antibodies. Method 1 was a two-step sandwich assay with polyclonal antibody as capture and europium chelate-labeled monoclonal antibody as tracer. Method 2 was a one-step heterogeneous competitive assay using immobilized polyclonal antibody and europium-labeled cystatin C. Method 3 was a one-step homogeneous competitive assay with europium-labeled polyclonal antibody as donor and cyanine 5-labeled cystatin C as acceptor. All three assays were evaluated with plasma samples and their performance was compared to a conventional particle-enhanced turbidimetric immunoassay (PETIA). Method 3 was the easiest to perform, with incubation at ambient temperature for 10 min and 20 mu L of sample, while methods 1 and 2 had washing steps, took 40 min and 15 min at 37 degrees C, respectively, but used only 10 mu L of 100- or 10-fold diluted sample, respectively. The working ranges for methods 1, 2 and 3 were 0.0005-0.2, 0.05-1.0 and 0.25-20 mg/L, respectively. Kinetics for method 3 was the fastest with >95% binding completion and for method 2 the slowest with 60% binding completion. All three methods showed good correlation to PETIA, but produced higher cystatin C levels than PETIA. Methods 1 and 3 offered the most favorable performance characteristics and especially method 3 enabled rapid and simple measurement of circulating cystatin C. (C) 2012 Elsevier B.V. All rights reserved.}},
author = {{Ristiniemi, Noora and Qin, Qiu-Ping and Lindström, Veronica and Grubb, Anders and Pettersson, Kim}},
issn = {{1872-7905}},
keywords = {{Cystatin C; Immunoassay; Heterogeneous assay; Homogeneous assay; Time-resolved fluorescence}},
language = {{eng}},
number = {{1-2}},
pages = {{56--61}},
publisher = {{Elsevier}},
series = {{Journal of Immunological Methods}},
title = {{Quantification of cystatin C by time-resolved fluorometry-based immunoassays}},
url = {{http://dx.doi.org/10.1016/j.jim.2012.02.004}},
doi = {{10.1016/j.jim.2012.02.004}},
volume = {{378}},
year = {{2012}},
}