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Development of an efficient extraction methodology to analyse potential inflammatory biomarkers from sebum

Development of an efficient extraction methodology to analyse potential inflammatory biomarkers from sebum
Development of an efficient extraction methodology to analyse potential inflammatory biomarkers from sebum
Introduction: proteins, such as cytokines and chemokines, are present in varying concentrations in a range of biofluids, with an important signalling role in maintaining homeostasis. Commercial tapes have been employed to non-invasively collect these potential biomarkers in sebum from the skin surface to examine their concentrations in conditions including acne, atopic dermatitis and pressure ulcers. However, the identification of robust biomarker candidates is limited by the low abundance of specific proteins extracted by current methodologies. Therefore, this study was designed to develop an optimized extraction method for potential inflammatory biomarkers in sebum collected with Sebutapes.

Methods: commercial tapes (Sebutapes) coated with synthetic sebum were used to systematically evaluate the effects of chemical and mechanical stimuli on extraction efficiency. Varying concentrations of high and low abundance biomarkers (IL-1α, IL-6, Il-8, INF-γ, TNF-α and IL-1RA) were used to spike the synthetic sebum samples. Methodological variables included different surfactants, mechanical stimuli and buffer volume. Extraction efficiency was estimated using immunoassay kits from the extracted buffer.

Results: the results revealed that the use of a surfactant, i.e. β-dodecyl maltoside in addition to the mechanical stimuli, namely sonication and centrifugation resulted in an increased recovery of cytokines, ranging from 80% for high-abundant cytokines, such as IL-1α and IL-1RA, and up to 50% for low-abundance cytokines, including TNF-alpha, IL-6 and IL-8. Compared to previous methods, the new extraction protocol resulted in between an 1.5 - 2.0 fold increase in extraction efficiency.

Conclusion: the study revealed that there was a high degree of variability in the extraction efficiency of different cytokines. However, improved efficiency was achieved across all cytokines with selective surfactants and mechanical stimuli. The optimised protocol will provide means to detect low levels of potential biomarkers from skin surface, enabling the evaluation of local changes in pro- and anti-inflammatory cytokines present in different skin conditions.
1660-5527
Jayabal, Hemalatha
8f2b053c-b614-4af2-b332-8ee861ab75f6
Bader, Daniel
9884d4f6-2607-4d48-bf0c-62bdcc0d1dbf
Worsley, Peter
6d33aee3-ef43-468d-aef6-86d190de6756
Jayabal, Hemalatha
8f2b053c-b614-4af2-b332-8ee861ab75f6
Bader, Daniel
9884d4f6-2607-4d48-bf0c-62bdcc0d1dbf
Worsley, Peter
6d33aee3-ef43-468d-aef6-86d190de6756

Jayabal, Hemalatha, Bader, Daniel and Worsley, Peter (2022) Development of an efficient extraction methodology to analyse potential inflammatory biomarkers from sebum. Skin Pharmacoloy and Physiology. (doi:10.1159/000528653).

Record type: Article

Abstract

Introduction: proteins, such as cytokines and chemokines, are present in varying concentrations in a range of biofluids, with an important signalling role in maintaining homeostasis. Commercial tapes have been employed to non-invasively collect these potential biomarkers in sebum from the skin surface to examine their concentrations in conditions including acne, atopic dermatitis and pressure ulcers. However, the identification of robust biomarker candidates is limited by the low abundance of specific proteins extracted by current methodologies. Therefore, this study was designed to develop an optimized extraction method for potential inflammatory biomarkers in sebum collected with Sebutapes.

Methods: commercial tapes (Sebutapes) coated with synthetic sebum were used to systematically evaluate the effects of chemical and mechanical stimuli on extraction efficiency. Varying concentrations of high and low abundance biomarkers (IL-1α, IL-6, Il-8, INF-γ, TNF-α and IL-1RA) were used to spike the synthetic sebum samples. Methodological variables included different surfactants, mechanical stimuli and buffer volume. Extraction efficiency was estimated using immunoassay kits from the extracted buffer.

Results: the results revealed that the use of a surfactant, i.e. β-dodecyl maltoside in addition to the mechanical stimuli, namely sonication and centrifugation resulted in an increased recovery of cytokines, ranging from 80% for high-abundant cytokines, such as IL-1α and IL-1RA, and up to 50% for low-abundance cytokines, including TNF-alpha, IL-6 and IL-8. Compared to previous methods, the new extraction protocol resulted in between an 1.5 - 2.0 fold increase in extraction efficiency.

Conclusion: the study revealed that there was a high degree of variability in the extraction efficiency of different cytokines. However, improved efficiency was achieved across all cytokines with selective surfactants and mechanical stimuli. The optimised protocol will provide means to detect low levels of potential biomarkers from skin surface, enabling the evaluation of local changes in pro- and anti-inflammatory cytokines present in different skin conditions.

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Accepted/In Press date: 5 December 2022
Published date: 2022

Identifiers

Local EPrints ID: 473627
URI: http://eprints.soton.ac.uk/id/eprint/473627
ISSN: 1660-5527
PURE UUID: c25f3cc4-9168-449c-a8b3-522e01d5e425
ORCID for Hemalatha Jayabal: ORCID iD orcid.org/0000-0003-4192-4464
ORCID for Daniel Bader: ORCID iD orcid.org/0000-0002-1208-3507
ORCID for Peter Worsley: ORCID iD orcid.org/0000-0003-0145-5042

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Date deposited: 25 Jan 2023 17:41
Last modified: 17 Mar 2024 03:15

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Contributors

Author: Hemalatha Jayabal ORCID iD
Author: Daniel Bader ORCID iD
Author: Peter Worsley ORCID iD

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