Pyronin Y (Pyronine G)

Flow Cytometric Detection of G0 in Live Cells by Hoechst 33342 and Pyronin Y Staining

Methods Mol Biol 2018;1686:49-57.29030811 10.1007/978-1-4939-7371-2_3

Hoechst 33342 and Pyronin Y double staining can be used to measure DNA and RNA content in live cells by flow cytometry. Quiescent cells at G0 phase have the same amount of DNA as cells at G1 phase but lower RNA levels compared to proliferating cells. Therefore, resting cells in G0 phase can be distinguished from proliferating cells in G1, S, and G2 M phases. This chapter describes a protocol for double staining of live cells with Hoechst 33342 and Pyronin Y. Combined with immunophenotyping of intact and live cells Hoechst 33342 and Pyronin Y staining is a powerful noninvasive method for the analysis and isolation of quiescent cells from any defined cell population.

Pyronin Y in the methyl-green-pyronin histological stain

Stain Technol 1955 Sep;30(5):213-30.13256168 10.3109/10520295509114469

Photophysics and photodynamics of Pyronin Y in n-alcohols

Luminescence 2018 Dec;33(8):1394-1400.30403000 10.1002/bio.3560

The photophysical properties and photodynamics of Pyronin Y (PyY) dye compound in seven polar protic solvents (n-alcohols) were examined as a function of temperature by using UV-visible, steady-state and time-resolved fluorescence spectroscopy techniques. To understand dye-solvent interactions, photophysical parameters including Stokes' shifts, fluorescence quantum yields and fluorescence lifetimes were determined. To examine the effect of solvent polarity, the difference between the ground state dipole moment and the excited state dipole moment was determined. For this purpose, the multiple regression analysis and the Kamlet-Taft technique were used. Moreover, photodynamic parameters, rotational relaxation times and steady-state anisotropy were calculated. The result showed that the specific interactions of PyY with the solvent molecules take place through hydrogen bonding. As the hydrocarbon chain of the alcohols gets longer, photophysical parameters diminish, probably because of weaker hydrogen bonding. Furthermore, it was found out that the dipole moment of excited states (μe ) is higher than that of the ground state (μg ). In addition, Brownian motions increased with the increasing temperature that weakened the fluorescence character of PyY. It was also revealed that the rotation of PyY increased with a prolonged hydrocarbon chain of alcohol series, due to the lesser extent of hydrogen bonding.

Application of Pyronin Y(G) in cytochemistry of nucleic acids

Cytometry 1987 Mar;8(2):138-45.2438101 10.1002/cyto.990080206

Chinese hamster ovary (CHO) cells or isolated nuclei were stained with Pyronin Y(PY) and analyzed by absorption or fluorescence microscopy, as well as by flow cytometry. Specificity of the staining reaction was assayed by testing sensitivity of the stainable material to RNase or DNase. The colored complexes detected by light absorption in fixed cells stained with PY are nonfluorescent and are most likely the products of condensation of single-stranded (ss) RNA by PY; the poly(rA) and poly(rA,rG) are the most sensitive to condensation. The products of PY interaction with double-stranded (ds) nucleic acids are fluorescent and can be detected in cells by cytofluorometry. PY used alone stains both DNA and RNA, and the staining capabilities of these nucleic acids vary depending upon the PY concentration at equilibrium; at a concentration above 330 microM, the RNA stainability decreases, perhaps due to its denaturation and condensation caused by the dye. In the presence of Hoechst 33342, PY can specifically stain RNA in fixed cells or isolated cell nuclei. Because only complexes of PY with ds RNA are fluorescent, this dye can be used as a probe of RNA conformation, e.g., to monitor denaturation of RNA in situ. The RNA stainability of mitotic cells is about 25% lower than that of cells in G2 phase, which indicates that during mitosis proportionately less cellular RNA is in the ds conformation. The advantages and limitations of the two cytochemical methods for DNA/RNA detection, one based on the use of Hoechst 33342 and PY, and another employing the metachromatic properties of acridine orange, are compared.

Spectroscopic investigations on the binding of Pyronin Y to human serum albumin

J Biomol Struct Dyn 2017 Jan;35(1):8-16.26646531 10.1080/07391102.2015.1128357

The interaction of Pyronin Y with human serum albumin (HSA) has been investigated systematically by fluorescence, absorption, fluorescence decay lifetime measurements, FTIR, synchronous fluorescence spectroscopy, and molecular modeling method. The spectroscopic and fluorescence quenching experiments show that Pyronin Y may show a static quenching mechanism with HSA. The specific binding distance of 1.96 nm between HSA and Pyronin Y was obtained via Förster non-radiation energy transfer method. The thermodynamic parameters indicate that the electrostatic interactions play a significant role during the binding process. In addition, synchronous fluorescence and FT-IR spectra indicated that the conformation and microenvironment of HSA were not influenced with the addition of Pyronin Y. The obtained results can be of biological significance in photodynamic therapy.