TY  - THES
AU  - Pismestrović, Marina
PY  - 2023
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/6138
AB  - In vitro metode koje simuliraju procese digestije se široko koriste za proučavanje 
gastrointestinalnog varenja hrane ili farmaceutskih proizvoda. Iako se nutricionističke studije još 
uvek smatraju “zlatnim standardom” za rešavanje pitanja vezanih za ishranu, in vitro metode imaju 
prednost zato što su brže, jeftinije, manje invazivne i nemaju etička ograničenja. Ovakve prednosti 
omogućavaju dalje skrining većeg broja uzoraka paralelno. Takođe, reproducibilnost, 
kontrolisanost uslova i lako uzorkovanje čine in vitro modele veoma pogodnim za istraživanja i 
postavljanje hipoteza. 
U ovom radu ispitana je kinetika ekstrakcije ukupnih proteina sirovih i kuvanih zapadno-pacifičkih 
kozica (lat. Solenocera melantho) u različitim fiziološki relevantnim tečnostima. Količina i 
elektroforetski profil ekstrahovanih ukupnih proteina upoređeni su vizuelno pomoću SDS PAGE 
i histogramski nakon Bredfordove metode za određivanje ukupne koncentracije proteina u datim 
alikvotima. Prateći in vitro standardizovani statički protokol tzv. INFOGEST urađene su bile tri 
različite digestije sirovih i kuvanih kozica. Imunogenost peptidnih fragmenata dobijenih u 
različitim fazama digestije ispitana je Western blotom korišćenjem poliklonskih anti-TPM antitela
i seruma osoba alergičnih na morske plodove i grinje. Talozi zaostali nakon završene gastrične i 
intestinalne faze zasebno su ispitani u slučaju da neki od imunogenih peptidnih fragmenata pada 
u talog nakon/tokom centrifugiranja pre spremanja uzorka za elektroforezu. Finalno, urađena je 
ELISA zarad kvantifikacije TPM-a u ekstraktima sirovih kozica u simuliranim fiziološkim 
tečnostima, kao i alikvotima uzetih tokom in vitro digestije nakon završene gastrične i intestinalne
faze.
T1  - Analiza imunoreaktivnosti peptida i proteina  kozica dobijenih u simuliranim uslovima  gastrointestinalne digestije korišćenjem  INFOGEST protokola
SP  - 2
EP  - 76
UR  - https://hdl.handle.net/21.15107/rcub_cherry_6138
ER  - 

@mastersthesis{
author = "Pismestrović, Marina",
year = "2023",
abstract = "In vitro metode koje simuliraju procese digestije se široko koriste za proučavanje 
gastrointestinalnog varenja hrane ili farmaceutskih proizvoda. Iako se nutricionističke studije još 
uvek smatraju “zlatnim standardom” za rešavanje pitanja vezanih za ishranu, in vitro metode imaju 
prednost zato što su brže, jeftinije, manje invazivne i nemaju etička ograničenja. Ovakve prednosti 
omogućavaju dalje skrining većeg broja uzoraka paralelno. Takođe, reproducibilnost, 
kontrolisanost uslova i lako uzorkovanje čine in vitro modele veoma pogodnim za istraživanja i 
postavljanje hipoteza. 
U ovom radu ispitana je kinetika ekstrakcije ukupnih proteina sirovih i kuvanih zapadno-pacifičkih 
kozica (lat. Solenocera melantho) u različitim fiziološki relevantnim tečnostima. Količina i 
elektroforetski profil ekstrahovanih ukupnih proteina upoređeni su vizuelno pomoću SDS PAGE 
i histogramski nakon Bredfordove metode za određivanje ukupne koncentracije proteina u datim 
alikvotima. Prateći in vitro standardizovani statički protokol tzv. INFOGEST urađene su bile tri 
različite digestije sirovih i kuvanih kozica. Imunogenost peptidnih fragmenata dobijenih u 
različitim fazama digestije ispitana je Western blotom korišćenjem poliklonskih anti-TPM antitela
i seruma osoba alergičnih na morske plodove i grinje. Talozi zaostali nakon završene gastrične i 
intestinalne faze zasebno su ispitani u slučaju da neki od imunogenih peptidnih fragmenata pada 
u talog nakon/tokom centrifugiranja pre spremanja uzorka za elektroforezu. Finalno, urađena je 
ELISA zarad kvantifikacije TPM-a u ekstraktima sirovih kozica u simuliranim fiziološkim 
tečnostima, kao i alikvotima uzetih tokom in vitro digestije nakon završene gastrične i intestinalne
faze.",
title = "Analiza imunoreaktivnosti peptida i proteina  kozica dobijenih u simuliranim uslovima  gastrointestinalne digestije korišćenjem  INFOGEST protokola",
pages = "2-76",
url = "https://hdl.handle.net/21.15107/rcub_cherry_6138"
}

Pismestrović, M.. (2023). Analiza imunoreaktivnosti peptida i proteina  kozica dobijenih u simuliranim uslovima  gastrointestinalne digestije korišćenjem  INFOGEST protokola. , 2-76.
https://hdl.handle.net/21.15107/rcub_cherry_6138

Pismestrović M. Analiza imunoreaktivnosti peptida i proteina  kozica dobijenih u simuliranim uslovima  gastrointestinalne digestije korišćenjem  INFOGEST protokola. 2023;:2-76.
https://hdl.handle.net/21.15107/rcub_cherry_6138 .

Pismestrović, Marina, "Analiza imunoreaktivnosti peptida i proteina  kozica dobijenih u simuliranim uslovima  gastrointestinalne digestije korišćenjem  INFOGEST protokola" (2023):2-76,
https://hdl.handle.net/21.15107/rcub_cherry_6138 .

TY  - CONF
AU  - Pismestrović, Marina
AU  - Radomirović, Mirjana Ž.
AU  - Čolaković, Maša
AU  - Ćirković-Veličković, Tanja
PY  - 2022
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/6026
AB  - Tropomyosin (TPM) is a major allergen among different shellfish species. Developing sensitive, specific, and reliable methods for quantifying TPM in food products is crucial for persons allergic to shellfish. Commonly used extraction buffers often show shortcomings in their extraction efficiency, which is why sometimes the presence of some allergens can be overlooked in the biological material. Therefore, this work aimed to optimize Mediterranean mussel TPM extraction conditions and develop a sandwich ELISA method for TPM quantification. Several extraction buffers were tested for their efficiency in recovering proteins from fresh frozen and cooked mussels during 2 and 24 hours of extraction. The protein content was quantified using the Bradford protein assay. Protein components of soluble extracts were profiled using SDS-PAGE. TPM presence in soluble extracts was confirmed by Western blot using both monoclonal and polyclonal anti-TPM antibodies. Sandwich ELISA was developed and used to quantify TPM content. None of the extraction buffers showed a significant difference in total protein content between 2 and 24 hours of extraction, indicating that 2 hours is sufficient for protein recovery in both raw and cooked mussels. Significantly fewer proteins were extracted from cooked mussels compared to raw mussels. Densitometrically estimated TPM concentrations indicate that PBS containing 1M NaCl (PBSN) extracts around 40% more TPM than PBS. Carbonate buffers extract even three times higher amounts of TPM than traditionally used extraction buffer PBS. Developed sandwich ELISA has shown not to be reliable for quantifying TPM from mussels, significantly underestimating its concentration, as concluded by comparing TPM concentrations obtained by ELISA with those obtained densitometrically. Therefore, Western blot has been used as an alternative method for mussel TPM quantification. The linear range for TPM quantification using Western blot was between 1.25 and 10 µg/ml. TPM concentrations in mussel extracts estimated using Western blot correlated well with those calculated by densitometric gel analysis. Further work will be aimed at improving the sensitivity of the presented methods and developing new methods for TPM quantification.
PB  - Beograd : Srpsko hemijsko društvo
PB  - Beograd : Klub mladih hemičara Srbije
C3  - 8th Conference of Young Chemists of Serbia, Belgrade, Serbia, 29th October, 2022. In: Book of Abstracts
T1  - Optimization of extraction conditions of tropomyosin from Mediterranean mussel and its quantification by developed ELISA
SP  - 54
EP  - 54
UR  - https://hdl.handle.net/21.15107/rcub_cherry_6026
ER  - 

@conference{
author = "Pismestrović, Marina and Radomirović, Mirjana Ž. and Čolaković, Maša and Ćirković-Veličković, Tanja",
year = "2022",
abstract = "Tropomyosin (TPM) is a major allergen among different shellfish species. Developing sensitive, specific, and reliable methods for quantifying TPM in food products is crucial for persons allergic to shellfish. Commonly used extraction buffers often show shortcomings in their extraction efficiency, which is why sometimes the presence of some allergens can be overlooked in the biological material. Therefore, this work aimed to optimize Mediterranean mussel TPM extraction conditions and develop a sandwich ELISA method for TPM quantification. Several extraction buffers were tested for their efficiency in recovering proteins from fresh frozen and cooked mussels during 2 and 24 hours of extraction. The protein content was quantified using the Bradford protein assay. Protein components of soluble extracts were profiled using SDS-PAGE. TPM presence in soluble extracts was confirmed by Western blot using both monoclonal and polyclonal anti-TPM antibodies. Sandwich ELISA was developed and used to quantify TPM content. None of the extraction buffers showed a significant difference in total protein content between 2 and 24 hours of extraction, indicating that 2 hours is sufficient for protein recovery in both raw and cooked mussels. Significantly fewer proteins were extracted from cooked mussels compared to raw mussels. Densitometrically estimated TPM concentrations indicate that PBS containing 1M NaCl (PBSN) extracts around 40% more TPM than PBS. Carbonate buffers extract even three times higher amounts of TPM than traditionally used extraction buffer PBS. Developed sandwich ELISA has shown not to be reliable for quantifying TPM from mussels, significantly underestimating its concentration, as concluded by comparing TPM concentrations obtained by ELISA with those obtained densitometrically. Therefore, Western blot has been used as an alternative method for mussel TPM quantification. The linear range for TPM quantification using Western blot was between 1.25 and 10 µg/ml. TPM concentrations in mussel extracts estimated using Western blot correlated well with those calculated by densitometric gel analysis. Further work will be aimed at improving the sensitivity of the presented methods and developing new methods for TPM quantification.",
publisher = "Beograd : Srpsko hemijsko društvo, Beograd : Klub mladih hemičara Srbije",
journal = "8th Conference of Young Chemists of Serbia, Belgrade, Serbia, 29th October, 2022. In: Book of Abstracts",
title = "Optimization of extraction conditions of tropomyosin from Mediterranean mussel and its quantification by developed ELISA",
pages = "54-54",
url = "https://hdl.handle.net/21.15107/rcub_cherry_6026"
}

Pismestrović, M., Radomirović, M. Ž., Čolaković, M.,& Ćirković-Veličković, T.. (2022). Optimization of extraction conditions of tropomyosin from Mediterranean mussel and its quantification by developed ELISA. in 8th Conference of Young Chemists of Serbia, Belgrade, Serbia, 29th October, 2022. In: Book of Abstracts
Beograd : Srpsko hemijsko društvo., 54-54.
https://hdl.handle.net/21.15107/rcub_cherry_6026

Pismestrović M, Radomirović MŽ, Čolaković M, Ćirković-Veličković T. Optimization of extraction conditions of tropomyosin from Mediterranean mussel and its quantification by developed ELISA. in 8th Conference of Young Chemists of Serbia, Belgrade, Serbia, 29th October, 2022. In: Book of Abstracts. 2022;:54-54.
https://hdl.handle.net/21.15107/rcub_cherry_6026 .

Pismestrović, Marina, Radomirović, Mirjana Ž., Čolaković, Maša, Ćirković-Veličković, Tanja, "Optimization of extraction conditions of tropomyosin from Mediterranean mussel and its quantification by developed ELISA" in 8th Conference of Young Chemists of Serbia, Belgrade, Serbia, 29th October, 2022. In: Book of Abstracts (2022):54-54,
https://hdl.handle.net/21.15107/rcub_cherry_6026 .

TY  - CONF
AU  - Radomirović, Mirjana Ž.
AU  - Čolaković, Maša
AU  - Pismestrović, Marina
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Rajković, Andreja
AU  - Ćirković-Veličković, Tanja
PY  - 2022
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/6027
AB  - Tropomyosin (TPM) is considered a major allergen among different shellfish species. Therefore, the development of methods for quantifying TPM in food products is crucial for allergic persons. Several extraction buffers were tested for their efficiency in recovering proteins from fresh frozen and cooked razor mud shrimp during 2 and 24 hours of extraction. The protein content was quantified using the Bradford protein assay. SDS-PAGE was used for protein profiling of soluble extracts. Sandwich ELISA was developed and used to quantify TPM content. None of the extraction buffers showed a significant difference in total protein content between 2 and 24 hours of extraction. Significantly fewer proteins were extracted from cooked shrimp compared to the raw shrimp. ELISA quantification showed that phosphate-buffered saline (PBS) containing 1 M NaCl (PBSN) and carbonate buffer, pH 10, extracted approximately 6 times higher amount of tropomyosin in comparison to PBS, highlighting the importance of choosing the appropriate extraction buffer for the precise quantification of TPM. Traditionally used extraction buffer PBS could significantly underestimate shrimp TPM content.
PB  - Beograd : Srpsko hemijsko društvo
C3  - 58th Meeting of the Serbian Chemical Society, Belgrade, Serbia, 9th-10th June, 2022. In: Book of Abstracts and Proceedings
T1  - Optimization of extraction conditions of tropomyosin from razor mud shrimp and its quantification by developed ELISA
SP  - 64
EP  - 64
UR  - https://hdl.handle.net/21.15107/rcub_cherry_6027
ER  - 

@conference{
author = "Radomirović, Mirjana Ž. and Čolaković, Maša and Pismestrović, Marina and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković-Veličković, Tanja",
year = "2022",
abstract = "Tropomyosin (TPM) is considered a major allergen among different shellfish species. Therefore, the development of methods for quantifying TPM in food products is crucial for allergic persons. Several extraction buffers were tested for their efficiency in recovering proteins from fresh frozen and cooked razor mud shrimp during 2 and 24 hours of extraction. The protein content was quantified using the Bradford protein assay. SDS-PAGE was used for protein profiling of soluble extracts. Sandwich ELISA was developed and used to quantify TPM content. None of the extraction buffers showed a significant difference in total protein content between 2 and 24 hours of extraction. Significantly fewer proteins were extracted from cooked shrimp compared to the raw shrimp. ELISA quantification showed that phosphate-buffered saline (PBS) containing 1 M NaCl (PBSN) and carbonate buffer, pH 10, extracted approximately 6 times higher amount of tropomyosin in comparison to PBS, highlighting the importance of choosing the appropriate extraction buffer for the precise quantification of TPM. Traditionally used extraction buffer PBS could significantly underestimate shrimp TPM content.",
publisher = "Beograd : Srpsko hemijsko društvo",
journal = "58th Meeting of the Serbian Chemical Society, Belgrade, Serbia, 9th-10th June, 2022. In: Book of Abstracts and Proceedings",
title = "Optimization of extraction conditions of tropomyosin from razor mud shrimp and its quantification by developed ELISA",
pages = "64-64",
url = "https://hdl.handle.net/21.15107/rcub_cherry_6027"
}

Radomirović, M. Ž., Čolaković, M., Pismestrović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković-Veličković, T.. (2022). Optimization of extraction conditions of tropomyosin from razor mud shrimp and its quantification by developed ELISA. in 58th Meeting of the Serbian Chemical Society, Belgrade, Serbia, 9th-10th June, 2022. In: Book of Abstracts and Proceedings
Beograd : Srpsko hemijsko društvo., 64-64.
https://hdl.handle.net/21.15107/rcub_cherry_6027

Radomirović MŽ, Čolaković M, Pismestrović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković-Veličković T. Optimization of extraction conditions of tropomyosin from razor mud shrimp and its quantification by developed ELISA. in 58th Meeting of the Serbian Chemical Society, Belgrade, Serbia, 9th-10th June, 2022. In: Book of Abstracts and Proceedings. 2022;:64-64.
https://hdl.handle.net/21.15107/rcub_cherry_6027 .

Radomirović, Mirjana Ž., Čolaković, Maša, Pismestrović, Marina, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković-Veličković, Tanja, "Optimization of extraction conditions of tropomyosin from razor mud shrimp and its quantification by developed ELISA" in 58th Meeting of the Serbian Chemical Society, Belgrade, Serbia, 9th-10th June, 2022. In: Book of Abstracts and Proceedings (2022):64-64,
https://hdl.handle.net/21.15107/rcub_cherry_6027 .

TY  - THES
AU  - Pismestrović, Marina
PY  - 2022
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/5483
AB  - Tropomiozini su visoko očuvani kontraktilni proteini kod kičmenjaka i beskičmenjaka, molekulske mase od oko 35-38 kDa. Uprkos njihovoj relativno maloj zastupljenosti, kod porodice beskičmenjaka prepoznati su kao pan-alergeni. Prema statistici tropomiozini su treći najčešći uzrok anafilakse izazvane hranom. U ovom radu ispitani su uslovi optimalne ekstrakcije proteina sirovih i kuvanih dagnji. Krajnji cilj je bila kvantifikacija tropomiozina u dobijenim ekstraktima sirovih i kuvanih dagnji, razvijanjem i optimizacijom uslova ELISA testa.
Za potrebe optimizacije uslova ekstrakcije korišćeno je nekoliko pufera različitih pH vrednosti i jonske sile, kao i različito vreme ekstrakcije (2 h i 24 h). Kao model organizam izabrana je mediteranska dagnja. Kvantifikacija proteina u ekstraktima urađena je Bredfordovom metodom, proteinski profili ekstrakata vizualizovani su pomoću natrijum-dodecilsulfat poliakrilamidne gel elektroforeze (SDS-PAGE), a potom je potvrđeno prisustvo tropomiozina Western blotom sa monoklonskim i poliklonskim antitelima. Nakon optimizacije ELISA testa urađena je kvantifikacija tropomiozina u dobijenim ekstraktima.
Prema dobijenim rezultatima, nije primećena statistički značajna razlika u koncentracijama proteina u ekstraktima dobijenim nakon 2 h i 24 h. Ekstrakcija proteina dagnji pokazala se efikasnijom sa porastom pH pufera. Vrednosti koncentracija proteina u ekstraktima sirovih i kuvanih dagnji, dobijene Bredfordovom metodom, u skladu su sa elektroforetskim profilima ekstrakata. Denzitometrijom, sa gela kuvanih uzoraka nakon SDS-PAGE, procenjene su bile koncentracije tropomiozina. Western blotom, odrađenim nakon SDS-PAGE, potvrđeno je prisustvo tropomiozina. Pokazano je da tropomiozin kuvanih ekstrakata daje jači signal od tropomiozina iz sirovih solubilnih ekstrakata dagnji. Radi kvantifikacije tropomiozina razvijen je sendvič ELISA test i izračunati su limit detekcije i limit kvantifikacije testa. Koncentracije tropomiozina u ekstraktima dobijene pomoću ELISA testa bile su za tri reda veličine niže od koncentracija procenjenih denzitometrijom, što pokazuje da se razvijeni ELISA test, kao takav, iako specifičan, ne može koristiti za pouzdanu i preciznu kvantifikaciju tropomiozina dagnji. Kao dobro rešenje, za prevazilaženje ovih ograničenja koja se javljaju kod ELISA testa, pokazala se kvantifikacija Western blotom.
T1  - Optimizacija uslova ekstrakcije i kvantifikacija tropomiozina dagnji ELISA testom
SP  - 1
EP  - 52
UR  - https://hdl.handle.net/21.15107/rcub_cherry_5483
ER  - 

@misc{
author = "Pismestrović, Marina",
year = "2022",
abstract = "Tropomiozini su visoko očuvani kontraktilni proteini kod kičmenjaka i beskičmenjaka, molekulske mase od oko 35-38 kDa. Uprkos njihovoj relativno maloj zastupljenosti, kod porodice beskičmenjaka prepoznati su kao pan-alergeni. Prema statistici tropomiozini su treći najčešći uzrok anafilakse izazvane hranom. U ovom radu ispitani su uslovi optimalne ekstrakcije proteina sirovih i kuvanih dagnji. Krajnji cilj je bila kvantifikacija tropomiozina u dobijenim ekstraktima sirovih i kuvanih dagnji, razvijanjem i optimizacijom uslova ELISA testa.
Za potrebe optimizacije uslova ekstrakcije korišćeno je nekoliko pufera različitih pH vrednosti i jonske sile, kao i različito vreme ekstrakcije (2 h i 24 h). Kao model organizam izabrana je mediteranska dagnja. Kvantifikacija proteina u ekstraktima urađena je Bredfordovom metodom, proteinski profili ekstrakata vizualizovani su pomoću natrijum-dodecilsulfat poliakrilamidne gel elektroforeze (SDS-PAGE), a potom je potvrđeno prisustvo tropomiozina Western blotom sa monoklonskim i poliklonskim antitelima. Nakon optimizacije ELISA testa urađena je kvantifikacija tropomiozina u dobijenim ekstraktima.
Prema dobijenim rezultatima, nije primećena statistički značajna razlika u koncentracijama proteina u ekstraktima dobijenim nakon 2 h i 24 h. Ekstrakcija proteina dagnji pokazala se efikasnijom sa porastom pH pufera. Vrednosti koncentracija proteina u ekstraktima sirovih i kuvanih dagnji, dobijene Bredfordovom metodom, u skladu su sa elektroforetskim profilima ekstrakata. Denzitometrijom, sa gela kuvanih uzoraka nakon SDS-PAGE, procenjene su bile koncentracije tropomiozina. Western blotom, odrađenim nakon SDS-PAGE, potvrđeno je prisustvo tropomiozina. Pokazano je da tropomiozin kuvanih ekstrakata daje jači signal od tropomiozina iz sirovih solubilnih ekstrakata dagnji. Radi kvantifikacije tropomiozina razvijen je sendvič ELISA test i izračunati su limit detekcije i limit kvantifikacije testa. Koncentracije tropomiozina u ekstraktima dobijene pomoću ELISA testa bile su za tri reda veličine niže od koncentracija procenjenih denzitometrijom, što pokazuje da se razvijeni ELISA test, kao takav, iako specifičan, ne može koristiti za pouzdanu i preciznu kvantifikaciju tropomiozina dagnji. Kao dobro rešenje, za prevazilaženje ovih ograničenja koja se javljaju kod ELISA testa, pokazala se kvantifikacija Western blotom.",
title = "Optimizacija uslova ekstrakcije i kvantifikacija tropomiozina dagnji ELISA testom",
pages = "1-52",
url = "https://hdl.handle.net/21.15107/rcub_cherry_5483"
}

Pismestrović, M.. (2022). Optimizacija uslova ekstrakcije i kvantifikacija tropomiozina dagnji ELISA testom. , 1-52.
https://hdl.handle.net/21.15107/rcub_cherry_5483

Pismestrović M. Optimizacija uslova ekstrakcije i kvantifikacija tropomiozina dagnji ELISA testom. 2022;:1-52.
https://hdl.handle.net/21.15107/rcub_cherry_5483 .

Pismestrović, Marina, "Optimizacija uslova ekstrakcije i kvantifikacija tropomiozina dagnji ELISA testom" (2022):1-52,
https://hdl.handle.net/21.15107/rcub_cherry_5483 .