Sažetak | Fluorescencija je pojava u kojoj fluorescentna čestica apsorbira svjetlo određene valne duljine te prelazi u pobuđeno stanje. Dok se fluorescentna čestica nalazi u pobuđenom stanju, dio energije se prenosi na obližnje molekule ili se raspršuje u molekularnim sudarima. Na posljetku, preostala se energija emitira kao foton kako bi se elektron vratio u osnovno stanje. Fluorescentne molekule se mogu podijeliti u tri opće skupine, a to su: biološki fluorofori, kvantne točke te organske boje. Navedene fluorescentne molekule se najčešće koriste prilikom fluorescentnog obilježavanja. Naime, radi se o postupku u kojem se fluorofor kovalentno veže na drugu molekulu, kao što je nukleinska kiselina ili protein. Obično se to postiže koristeći reaktivni derivat fluorofora koji se selektivno veže na funkcionalnu skupinu prisutnu u ciljanoj molekuli. U ovom radu se govori o fluorescentnom označavanju proteina pomoću organskih boja koje se vrši tako da se funkcionalna skupina određene fluorescentne boje veže na odgovarajuću funkcionalnu skupinu proteina, a najčešće su to: primarni amini, karboksilna skupina, sulfhidrilna skupina te karbonilna skupina. Nadalje, u ovom radu se također govori o funkcionalnim skupinama koje se najčešće koriste u službi obilježavanja proteina. Navedeni su primjeri najpoznatijih fluorescentnih boja te je sintetizirana boja 2,8- dietil-5-(4-karboksifenil)-1,3,7,9-tetrametildipirometen (BODIPY-COOH) (1) te njezin reaktivni derivat 5-(4-(N-sukcinimidoksikarbonil) fenil)-1,3,7,9-tetrametil-BODIPY (2). Sintetizirana boja (2) je najčešće korištena boja za konjugaciju sa strukturama koje sadržavaju amine stoga je od velikog značaja u kontekstu biooznačavanja. |
Sažetak (engleski) | Fluorescence is a phenomenon in which a fluorophore passes into an excitation state by
absorbing a photon of the light of certain wavelength. While the fluorophore is in the
excitation state, part of the energy is transferred to the adjacent molecules or lost due to
molecular collisions. Ultimately, the molecule returns to its basic state, while emitting energy
as a photon. There are three general groups of fluorescent molecules: biological fluorophors,
quantum dots and organic dyes. The most common use of fluorescent molecules is in
fluorescent labeling. Namely, fluorescent labeling entails the covalent binding of a
fluorophore to another molecule, such as a nucleic acid or a protein. Generally, this can be
accomplished by using a reactive derivative of a fluorophore which is capable of selectively
bind to a functional group present in the molecule of interest. This paper discusses the
fluorescence labeling of proteins using organic dyes. The functional group of a certain
fluorescent dye is linked to the corresponding functional group in the protein, most often of
which are primary amines, carboxyl groups, sulfhydryl groups and carbonyl groups.
Furthermore, this work discusses the various functional groups which are most commonly
used in protein labeling. Also, in this work are listed some of the examples of the most widely
used fluorescent dyes, and the compound 5-(4-carboxyphenyl)-2,8-diethyl-1,3,7,9-
tetramethyldipyrimomethene (BODIPY-COOH) (1) and its reactive derivative 5-(Nsuccinimidoxycarbonyl)
phenyl)-1,3,7,9-tetramethyl-BODIPY (2) have been synthesized. The
synthesized dye (2) is the most commonly used dye for conjugation with amino-containing
structures, therefore it is of great importance in the context of bio-labeling. |