Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR
Аутори
Radomirović, MirjanaGligorijević, Nikola
Stanić-Vučinić, Dragana
Rajković, Andreja
Ćirković Veličković, Tanja
Чланак у часопису (Објављена верзија)
Метаподаци
Приказ свих података о документуАпстракт
Tropomyosin is the major and predominant allergen among shellfish. This study developed
an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods.
The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs.
Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin
antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A
double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and
subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR
was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR
method is highly specific for the detection of crustacean tropomyosin and is highly precise in a
broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%.
Crustacean tropomyosin was also quantified in commercia...l food products. The reported immuno-
PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the
first immuno-PCR-based assay for the quantification of food allergen and food protein in general.
The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based
detection of traces of any food allergen that is currently being quantified with ELISA, which is of
critical importance for people with food allergies.
Кључне речи:
tropomyosin / immuno-PCR / crustacean allergen / ELISA / shellfish allergen / allergen quantificationИзвор:
International Journal of Molecular Sciences, 2023, 24, 20, 15410-Издавач:
- MDPI
Финансирање / пројекти:
- Science Fund of the Republic of Serbia, Program DIASPORA, #6504499, ShellPCR
- European Union’s Horizon 2020 research and innovation program under grant agreement No. 965173
- Министарство науке, технолошког развоја и иновација Републике Србије, институционално финансирање - 200168 (Универзитет у Београду, Хемијски факултет) (RS-MESTD-inst-2020-200168)
Институција/група
IHTMTY - JOUR AU - Radomirović, Mirjana AU - Gligorijević, Nikola AU - Stanić-Vučinić, Dragana AU - Rajković, Andreja AU - Ćirković Veličković, Tanja PY - 2023 UR - https://cer.ihtm.bg.ac.rs/handle/123456789/6782 AB - Tropomyosin is the major and predominant allergen among shellfish. This study developed an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods. The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs. Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR method is highly specific for the detection of crustacean tropomyosin and is highly precise in a broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%. Crustacean tropomyosin was also quantified in commercial food products. The reported immuno- PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the first immuno-PCR-based assay for the quantification of food allergen and food protein in general. The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based detection of traces of any food allergen that is currently being quantified with ELISA, which is of critical importance for people with food allergies. PB - MDPI T2 - International Journal of Molecular Sciences T1 - Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR VL - 24 IS - 20 SP - 15410 DO - 10.3390/ijms242015410 ER -
@article{ author = "Radomirović, Mirjana and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković Veličković, Tanja", year = "2023", abstract = "Tropomyosin is the major and predominant allergen among shellfish. This study developed an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods. The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs. Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR method is highly specific for the detection of crustacean tropomyosin and is highly precise in a broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%. Crustacean tropomyosin was also quantified in commercial food products. The reported immuno- PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the first immuno-PCR-based assay for the quantification of food allergen and food protein in general. The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based detection of traces of any food allergen that is currently being quantified with ELISA, which is of critical importance for people with food allergies.", publisher = "MDPI", journal = "International Journal of Molecular Sciences", title = "Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR", volume = "24", number = "20", pages = "15410", doi = "10.3390/ijms242015410" }
Radomirović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković Veličković, T.. (2023). Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR. in International Journal of Molecular Sciences MDPI., 24(20), 15410. https://doi.org/10.3390/ijms242015410
Radomirović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković Veličković T. Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR. in International Journal of Molecular Sciences. 2023;24(20):15410. doi:10.3390/ijms242015410 .
Radomirović, Mirjana, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković Veličković, Tanja, "Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR" in International Journal of Molecular Sciences, 24, no. 20 (2023):15410, https://doi.org/10.3390/ijms242015410 . .