Accurate quantification of β-hexosaminidase released from laboratory of allergic diseases 2 cells via liquid chromatography tandem mass spectrometry method

Lv, Yanni; Jia, Fu; Jia, Qi; Che, Delu; Lin, Yuanyuan; Han, Shengli; He, Langchong

doi:10.1016/j.chroma.2018.09.059

Cited by 10 publications

(7 citation statements)

References 34 publications

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“…We also measured the kinetics of TPS on the LCMS-8040 platform (Figure S3B) and by the substrate based method (Figure E). By referring to the kinetics results of HEXB in our previous work, we found that HEXB reached a maximum release amount at 30 min, whereas for TPS, the time point was 45 min. For the sake of convenience in the sample preparation process, we used the 45 min time point as a final incubation time.…”

Section: Resultsmentioning

confidence: 73%

“…After the 2-h incubation period at 37 °C, the OD values were read at 405 nm in a microplate reader. The substrate-based assay for determining released HEXB in LAD2 cells refers to our previous work …”

Section: Methodsmentioning

confidence: 99%

“…The substrate-based assay for determining released HEXB in LAD2 cells refers to our previous work. 27 In Solution Protein Digestion. The protein content in each supernatant sample was determined using the BCA assay, according to the manufacturer's instructions (Solarbio, Beijing, China).…”

Section: ■ Experimental Sectionmentioning

confidence: 99%

“…Optimization of Sensitivity and Simultaneous Measurement. In our previous work, 27 a mass spectrometry-based method was established for the determination of HEXB. Subsequently, we attempted to adapt this method for the TPS quantification, and the linear range and methodology were found to meet the requirements for quantification analysis (shown in Figure S3A and Table S1).…”

Section: T H I S C O N T E N T Imentioning

confidence: 99%

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Liquid Chromatography Tandem Mass Spectrometry Based Label-Free Quantification Method for Assessment of Allergen-Induced Anaphylactoid Reactions

Jia

et al. 2020

J. Am. Soc. Mass Spectrom.

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Mast cells are essential in mediating inflammatory processes. When activated, mast cells can rapidly release characteristic granules and various mediators into the interstitium. Tryptase (TPS) and β-hexosaminidase (HEXB) are typical protease mediators stored in granules and released upon activation. They have been recognized as important biomarkers of anaphylaxis, and the released level is associated with the severity of allergic reactions. In this study, a sensitive, accurate, and selective liquid chromatography tandem mass spectrometry (LC–MS/MS) method for simultaneously quantifying the two biomarkers was developed and validated in LAD2 cell culture supernatant, and P14R was used as internal standard. Good linearity was observed in the range of 50–2500 ng/mL for TPS and 10–2000 ng/mL for HEXB both with R 2 > 0.99. The matrix effect and recovery were both within acceptable limits. We quantified TPS and HEXB released from Laboratory of Allergic Disease 2 (LAD2) mast cells treated with several potential allergens, and the results demonstrate that the method can be used to investigate TPS and HEXB levels in LAD2 mast cell model during allergy research. We anticipate our approach to be a robust and sensitive assessment method for more biomarkers with similar kinetics characteristics and to be a major tool of allergic drug assessment or antiallergic drug development in research.

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Section: Resultsmentioning

confidence: 73%

Section: Methodsmentioning

confidence: 99%

Section: ■ Experimental Sectionmentioning

confidence: 99%

Section: T H I S C O N T E N T Imentioning

confidence: 99%

See 2 more Smart Citations

Liquid Chromatography Tandem Mass Spectrometry Based Label-Free Quantification Method for Assessment of Allergen-Induced Anaphylactoid Reactions

Jia

et al. 2020

J. Am. Soc. Mass Spectrom.

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“…β-Hexosaminidase, which is secreted from mast cells and basophils stimulated by antigens or degranulation inducers, is frequently considered as an indicator of anaphylactic reactions. 23) To investigate whether methanol extract of P. crocatum or compound 1 affects mast cell degranulation, we measured the amount of released β-hexosaminidase in the supernatants of RBL-2H3 cells stimulated by IgE-antigen complex. With the treatment of methanol extract or compound 1, the released β-hexosaminidase from RBL-2H3 cells was obviously reduced, and quercetin was used as a reference compound (Fig.…”

Section: Effect Of Methanol Extract and Compound 1 Of P Crocatum On β-Hexosaminidase Release In Rbl-2h3 Cellsmentioning

confidence: 99%

Anti-allergic Inflammatory Components from the Leaves of <i>Piper crocatum</i> Ruiz & Pav.

Gong

Guo

et al. 2021

Biological & Pharmaceutical Bulletin

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Piper crocatum Ruiz & Pav. (P. crocatum), a traditional medicinal plant, has been shown to possess various pharmacological activities, including anticancer activity, antioxidant activity, antibacterial activity, anti-hyperglycemic activity, anti-allergic inflammatory activity and others. To identify the potential anti-allergic inflammatory effective constituents of P. crocatum, 13 single compounds were isolated from the methanol extract of P. crocatum leaves, and their structures were identified by contrasting their NMR spectroscopic data and previously published papers. First, the anti-allergic inflammatory activities of these single compounds were examined by accessing immune function related biomarkers such as nitric oxide (NO) and β-hexosaminidase. We found that the methanol extract and catechaldehyde (compound 1) potently suppressed NO production. Additionally, Western blot analysis showed that P. crocatum methanol extract and compound 1 suppressed the production of NO by reducing inducible nitric oxide synthase (iNOS) expression in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. Consistent with these observations, P. crocatum methanol extract and compound 1 remarkably decreased β-hexosaminidase release from RBL-2H3 cells stimulated with 2,4-dinitrophenylated bovine serum albumin (DNP-BSA)-specific immunoglobulin E (IgE) antibodies. Furthermore, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay indicated that P. crocatum methanol extract and compound 1 exhibited no cytotoxicity to RAW264.7 and RBL-2H3 cells. Based on these findings, compound 1 is suggested as an active anti-allergic inflammatory component of P. crocatum.

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Thebaine induces anaphylactic reactions via the MRGPRX2 receptor pathway on mast cells

Lei

Liu

Ding

et al. 2022

Cellular Immunology

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Accurate quantification of β-hexosaminidase released from laboratory of allergic diseases 2 cells via liquid chromatography tandem mass spectrometry method

Cited by 10 publications

References 34 publications

Liquid Chromatography Tandem Mass Spectrometry Based Label-Free Quantification Method for Assessment of Allergen-Induced Anaphylactoid Reactions

Liquid Chromatography Tandem Mass Spectrometry Based Label-Free Quantification Method for Assessment of Allergen-Induced Anaphylactoid Reactions

Anti-allergic Inflammatory Components from the Leaves of <i>Piper crocatum</i> Ruiz & Pav.

Thebaine induces anaphylactic reactions via the MRGPRX2 receptor pathway on mast cells

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