ISO/TS 23366:2023
(Main)Nanotechnologies — Performance evaluation requirements for quantifying biomolecules using fluorescent nanoparticles in immunohistochemistry
Nanotechnologies — Performance evaluation requirements for quantifying biomolecules using fluorescent nanoparticles in immunohistochemistry
This document describes minimum requirements for performance evaluation of applying fluorescent nanoparticles in quantitative immuno-histochemistry.
Nanotechnologies — Exigences d'évaluation des performances pour la quantification de biomolécules en immunohistochimie à l'aide de nanoparticules fluorescentes
General Information
Standards Content (Sample)
TECHNICAL ISO/TS
SPECIFICATION 23366
First edition
2023-05
Nanotechnologies — Performance
evaluation requirements for
quantifying biomolecules using
fluorescent nanoparticles in
immunohistochemistry
Nanotechnologies — Exigences d'évaluation des performances pour
la quantification de biomolécules en immunohistochimie à l'aide de
nanoparticules fluorescentes
Reference number
© ISO 2023
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ii
Contents Page
Foreword .v
Introduction . vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 2
5 Selection of fluorescent nanoparticles . 3
5.1 General . 3
5.2 Characteristics of nanoparticle . 3
6 Quantification system .4
6.1 Overall design . 4
6.2 Antibody . 5
6.2.1 General . 5
6.2.2 Antibody type . 5
6.2.3 Target antigen . 5
6.2.4 Applicability of antibody to method . 6
6.2.5 Animal species . 6
6.2.6 Immunoglobulin class . 6
6.2.7 Titre . 6
6.2.8 Specificity . 7
6.3 Staining procedure . 7
6.3.1 Staining conditions . . 7
6.3.2 Robustness of staining procedure . 7
6.4 Image processing . 8
6.4.1 Image quality and relevant factors . 8
6.4.2 Selection of image processing software . 8
7 Comparability of results . 9
7.1 Light source adjustment. 9
7.2 Reference material . 9
8 Performance characteristics . 9
8.1 Background . 9
8.1.1 General . 9
8.1.2 Autofluorescence . 10
8.1.3 Nonspecific absorption of nanoparticles . 10
8.1.4 Nonspecific absorption of antibodies . 10
8.1.5 Loss of intensity and interferences . 10
8.2 Reference material dependent indices . 11
8.2.1 General . 11
8.2.2 Limit of detection (LOD) . 11
8.2.3 Limit of quantification (LOQ) . 11
8.2.4 Linearity and dynamic range . 11
8.3 Robustness .12
9 Validation and verification .12
9.1 General .12
9.2 Single lab precision .12
9.3 Reproducibility . 13
10 Reporting .13
Annex A (informative) Example of the reference material .14
iii
Annex B (informative) Example of the nanoparticle aggregation/agglomeration evaluation
method .16
Bibliography .18
iv
Foreword
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v
Introduction
Fluorescent nanoparticles are expanding their market into bio-technological and research fields as a
labelling material to be used for immunohistochemical staining.
Conventionally, various fluorescent dyes, including FITC (fluorescent isothiocyanate), rhodamine-
isothiocyanate and sulforhodamine 101 acid chloride, have been used for immunohistochemical
staining. They are still powerful tools for identifying localization of target biomolecules, for example,
proteins and sugar chains, mainly for qualitative analyses. They are also applied to quantitative analysis
in combination with various algorithms for calculating signal intensity related to the quantity of the
target biomolecules. The quantification system generally consists of sample preparation, staining,
microscopic observation and photography, and image processing for obtaining quantification results
as shown in Figure 1. For reliable measurement results of quantification, fluorescent dyes that are
brighter and more photostable by exposure to the excitation light are more appropriate.
Large number of fluorescent nanoparticles are available in the market. Generally, they show higher
brightness and are more resistant to photobleaching, compared to the conventional fluorescent dyes.
The characteristics of fluorescent nanoparticles can be an advantage for the quantification of target
biomolecules by immunohistochemical meth
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