Directional surface plasmon-coupled emission: Application for an immunoassay in whole blood

Evgenia G. Matveeva, Zygmunt Gryczynski, Joanna Malicka, Joanna Lukomska, Slawomir Makowiec, Klaus W. Berndt, Joseph R. Lakowicz, Ignacy Gryczynski

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

We present a new approach for performing fluorescence immunoassay in whole blood using fluorescently labeled anti-rabbit immunoglobulin G (IgG) on a silver surface. This approach, which is based on surface plasmon-coupled emission (SPCE), provides increased sensitivity and substantial background reduction due to exclusive selection of the signal from the fluorophores located near a bioaffinity surface. This article describes the effect of an optically dense sample matrix, namely human whole blood and serum, on the intensity of the SPCE. An antigen (rabbit IgG) was adsorbed to a slide covered with a thin silver metal layer, and the SPCE signal from the fluorophore-labeled anti-rabbit antibody, binding to the immobilized antigen, was detected. The effect of the sample matrix (buffer, human serum, or human whole blood) on the end-point immunoassay SPCE signal was studied. It was demonstrated that the kinetics of binding could be monitored directly in whole blood or serum. The results showed that human serum and human whole blood attenuate the SPCE end-point signal and the immunoassay kinetic signal only approximately two- and threefold, respectively, as compared with buffer, resulting in signals that are easily detectable even in whole blood. The high optical absorption of the hemoglobin can be tolerated because only fluorophores within a couple of hundred nanometers from the metallic film contribute to SPCE. Excited fluorophores outside the 200-nm layer do not contribute to SPCE, and their free space emission is not transmitted through the opaque metallic film into the glass substrate. We believe that SPCE has the potential of becoming a powerful approach for performing immunoassays based on surface-bound analytes or antibodies for many biomarkers directly in dense samples such as whole blood with no need for washing steps.

Original languageEnglish
Pages (from-to)161-167
Number of pages7
JournalAnalytical Biochemistry
Volume344
Issue number2
DOIs
StatePublished - 15 Sep 2005

Fingerprint

Immunoassay
Blood
Fluorophores
Rabbits
Serum
Silver
Buffers
Immunoglobulin G
Antigens
Metallic films
Glass
Anti-Idiotypic Antibodies
Hemoglobins
Biomarkers
Fluorescence
Metals
Antibodies
Kinetics
Washing
Light absorption

Keywords

  • Background suppression
  • Fluorescence immunoassay
  • Silver film
  • Surface plasmon-coupled emission
  • Whole blood

Cite this

Matveeva, Evgenia G. ; Gryczynski, Zygmunt ; Malicka, Joanna ; Lukomska, Joanna ; Makowiec, Slawomir ; Berndt, Klaus W. ; Lakowicz, Joseph R. ; Gryczynski, Ignacy. / Directional surface plasmon-coupled emission : Application for an immunoassay in whole blood. In: Analytical Biochemistry. 2005 ; Vol. 344, No. 2. pp. 161-167.
@article{0e9d3d2b2cff4f3c9c922c2602fadb00,
title = "Directional surface plasmon-coupled emission: Application for an immunoassay in whole blood",
abstract = "We present a new approach for performing fluorescence immunoassay in whole blood using fluorescently labeled anti-rabbit immunoglobulin G (IgG) on a silver surface. This approach, which is based on surface plasmon-coupled emission (SPCE), provides increased sensitivity and substantial background reduction due to exclusive selection of the signal from the fluorophores located near a bioaffinity surface. This article describes the effect of an optically dense sample matrix, namely human whole blood and serum, on the intensity of the SPCE. An antigen (rabbit IgG) was adsorbed to a slide covered with a thin silver metal layer, and the SPCE signal from the fluorophore-labeled anti-rabbit antibody, binding to the immobilized antigen, was detected. The effect of the sample matrix (buffer, human serum, or human whole blood) on the end-point immunoassay SPCE signal was studied. It was demonstrated that the kinetics of binding could be monitored directly in whole blood or serum. The results showed that human serum and human whole blood attenuate the SPCE end-point signal and the immunoassay kinetic signal only approximately two- and threefold, respectively, as compared with buffer, resulting in signals that are easily detectable even in whole blood. The high optical absorption of the hemoglobin can be tolerated because only fluorophores within a couple of hundred nanometers from the metallic film contribute to SPCE. Excited fluorophores outside the 200-nm layer do not contribute to SPCE, and their free space emission is not transmitted through the opaque metallic film into the glass substrate. We believe that SPCE has the potential of becoming a powerful approach for performing immunoassays based on surface-bound analytes or antibodies for many biomarkers directly in dense samples such as whole blood with no need for washing steps.",
keywords = "Background suppression, Fluorescence immunoassay, Silver film, Surface plasmon-coupled emission, Whole blood",
author = "Matveeva, {Evgenia G.} and Zygmunt Gryczynski and Joanna Malicka and Joanna Lukomska and Slawomir Makowiec and Berndt, {Klaus W.} and Lakowicz, {Joseph R.} and Ignacy Gryczynski",
year = "2005",
month = "9",
day = "15",
doi = "10.1016/j.ab.2005.07.005",
language = "English",
volume = "344",
pages = "161--167",
journal = "Analytical Biochemistry",
issn = "0003-2697",
publisher = "Academic Press Inc.",
number = "2",

}

Directional surface plasmon-coupled emission : Application for an immunoassay in whole blood. / Matveeva, Evgenia G.; Gryczynski, Zygmunt; Malicka, Joanna; Lukomska, Joanna; Makowiec, Slawomir; Berndt, Klaus W.; Lakowicz, Joseph R.; Gryczynski, Ignacy.

In: Analytical Biochemistry, Vol. 344, No. 2, 15.09.2005, p. 161-167.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Directional surface plasmon-coupled emission

T2 - Application for an immunoassay in whole blood

AU - Matveeva, Evgenia G.

AU - Gryczynski, Zygmunt

AU - Malicka, Joanna

AU - Lukomska, Joanna

AU - Makowiec, Slawomir

AU - Berndt, Klaus W.

AU - Lakowicz, Joseph R.

AU - Gryczynski, Ignacy

PY - 2005/9/15

Y1 - 2005/9/15

N2 - We present a new approach for performing fluorescence immunoassay in whole blood using fluorescently labeled anti-rabbit immunoglobulin G (IgG) on a silver surface. This approach, which is based on surface plasmon-coupled emission (SPCE), provides increased sensitivity and substantial background reduction due to exclusive selection of the signal from the fluorophores located near a bioaffinity surface. This article describes the effect of an optically dense sample matrix, namely human whole blood and serum, on the intensity of the SPCE. An antigen (rabbit IgG) was adsorbed to a slide covered with a thin silver metal layer, and the SPCE signal from the fluorophore-labeled anti-rabbit antibody, binding to the immobilized antigen, was detected. The effect of the sample matrix (buffer, human serum, or human whole blood) on the end-point immunoassay SPCE signal was studied. It was demonstrated that the kinetics of binding could be monitored directly in whole blood or serum. The results showed that human serum and human whole blood attenuate the SPCE end-point signal and the immunoassay kinetic signal only approximately two- and threefold, respectively, as compared with buffer, resulting in signals that are easily detectable even in whole blood. The high optical absorption of the hemoglobin can be tolerated because only fluorophores within a couple of hundred nanometers from the metallic film contribute to SPCE. Excited fluorophores outside the 200-nm layer do not contribute to SPCE, and their free space emission is not transmitted through the opaque metallic film into the glass substrate. We believe that SPCE has the potential of becoming a powerful approach for performing immunoassays based on surface-bound analytes or antibodies for many biomarkers directly in dense samples such as whole blood with no need for washing steps.

AB - We present a new approach for performing fluorescence immunoassay in whole blood using fluorescently labeled anti-rabbit immunoglobulin G (IgG) on a silver surface. This approach, which is based on surface plasmon-coupled emission (SPCE), provides increased sensitivity and substantial background reduction due to exclusive selection of the signal from the fluorophores located near a bioaffinity surface. This article describes the effect of an optically dense sample matrix, namely human whole blood and serum, on the intensity of the SPCE. An antigen (rabbit IgG) was adsorbed to a slide covered with a thin silver metal layer, and the SPCE signal from the fluorophore-labeled anti-rabbit antibody, binding to the immobilized antigen, was detected. The effect of the sample matrix (buffer, human serum, or human whole blood) on the end-point immunoassay SPCE signal was studied. It was demonstrated that the kinetics of binding could be monitored directly in whole blood or serum. The results showed that human serum and human whole blood attenuate the SPCE end-point signal and the immunoassay kinetic signal only approximately two- and threefold, respectively, as compared with buffer, resulting in signals that are easily detectable even in whole blood. The high optical absorption of the hemoglobin can be tolerated because only fluorophores within a couple of hundred nanometers from the metallic film contribute to SPCE. Excited fluorophores outside the 200-nm layer do not contribute to SPCE, and their free space emission is not transmitted through the opaque metallic film into the glass substrate. We believe that SPCE has the potential of becoming a powerful approach for performing immunoassays based on surface-bound analytes or antibodies for many biomarkers directly in dense samples such as whole blood with no need for washing steps.

KW - Background suppression

KW - Fluorescence immunoassay

KW - Silver film

KW - Surface plasmon-coupled emission

KW - Whole blood

UR - http://www.scopus.com/inward/record.url?scp=23944457867&partnerID=8YFLogxK

U2 - 10.1016/j.ab.2005.07.005

DO - 10.1016/j.ab.2005.07.005

M3 - Article

C2 - 16091280

AN - SCOPUS:23944457867

VL - 344

SP - 161

EP - 167

JO - Analytical Biochemistry

JF - Analytical Biochemistry

SN - 0003-2697

IS - 2

ER -