Malgorzata Lipowska

New Near-Infrared Cyanine Dyes for Labelling of Proteins

Abstract Isothiocyanato-functionalized cyanine dyes 7 and 11 for labelling of proteins at amino groups have been synthesized. The dyes and their adducts with amines show strong absorbance and fluorescence in the near-infrared region of 750-850 nm.

FACILE DERIVATIZATIONS OF HEPTAMETHINE CYANINE DYES

Abstract Treatment of dye 3 with sodium methoxide, morpholine, or diphenylphosphine gives the respective substituted derivatives 4, 5 and 6 in high yields. The reaction of 3 with sodium acetate followed by workup affords ketone 7 which is also obtained by demethylation of a methoxy derivative 4 under non-hydrolytic conditions.

Evaluation of near infrared dyes as labels for immunoassays utilizing laser diode detection : development of near infrared dye immunoassay (Nirdia)

Abstract Many recent studies of near infrared (NIR) dyes have shown their usefulness as probes for many analytical applications. Since these dyes have absorption maxima between 600 and 1000 nm, a region of low interference by most bioorganic molecules, their use as a label for biomolecules is practical. We report here the first study to evaluate the use of near infrared dyes as a quantitative label for immunoassays. A near infrared dye was derivatized with an isothiocyanate functional group and conjugated to goat anti-human immunoglobulins (GAHG). After purification by G-25 size exclusion chromatography, the conjugate was used in an immunoassay to detect and quantitate human immunoglobulins (Ig), the antigen for GAHG. Polystyrene microtiter plate wells were coated with varying amounts of human Ig. The Ig-coated wells were then exposed to an excess of NIRD-GAHG, and were subsequently read and quantitated by laser diode spectroscopy. An identical immunoassay was performed which utilized an enzyme(peroxidase...

Near-Infrared Heptamethine Cyanine Dyes: A New Tracer for Solid-Phase Immunoassays:

Near-infrared (near-IR) fluorescence has been used to develop a solid-phase immunoassay that detects trace amounts of human immunoglobulin (HuIgG). Various concentrations of HuIgG bound to a nitrocellulose surface were determined from the fluorescence generated by near-IR labeled goat anti-human antibody (GAHG) bound to the HuIgG. The GAHG was labeled with a heptamethine cyanine fluorophore that has spectral properties in the near-IR region (above 780 nm). These fluorophores are versatile because they can be modified for several bioanalytical applications. Fluorescence was detected with a near-IR fluorescence instrument previously developed in the laboratory. Two cyanine fluorophore labels were evaluated for the ability to selectively bind to GAHG on a nitrocellulose matrix with a minimal amount of background interference. After the most appropriate near-IR fluorophore was selected, the labeling of GAHG was optimized under aqueous conditions. The most effective GAHG–dye conjugates were used to develop an immunoassay to detect various concentrations of HuIgG. The results are presented, here. Solutions of HuIgG with concentrations as low as 10−10 molar have been detected with a minimum of interference.

Spectroscopic studies of a near-infrared absorbing pH sensitive aminodienone—carbocyanine dye system

Synthetic red and near-infrared absorbing dyes may be used as probe molecules in a large number of applications. Dyes exhibiting spectral changes with hydrogen ion concentration are useful as pH probes. Those dyes which have their absorption and fluorescence maxima in the long wavelength region of the visible spectral region are specially valuable because of decreased interference and semiconductor laser applications. In this paper we have evaluated an aminodienone dyes 1 which demostrates pH dependent absorption and fluorescence spectra as well as solvent polarity dependence. In organic solvents the long wavelength absorption band of the dye is in the reduced interference region. The absorption maximum is at 535 nm in neutral or alkaline solutions in methanol. The absorption spectra undergo a strong bathochromic shift in the presence of acids (lambda(max) = 709 nm) with a concomitant change in the fluorescence spectra. This pH sensitive dye was found to be specially especially useful for organic solvents. The analytical utility of this and similar near-infrared absorbing dyes is discussed.

Investigation of a near-infrared fiber optic immunosensor

The designs of NIR optical fiber immunosensors for the detection of biomolecules are discussed. The use of fiber optics combined with laser-excited fluorescence detection ((lambda) max equals 780 nm) and immunochemical techniques has provided the essential components for developing simpler and more practical, sensitive and selective immunosensors. The antibody is immobilized on the distal end of a surface-activated polymethyl methylacrylate sensor. As the probe is placed in different concentrations of antigenic substance, the antibody reacts with its corresponding antigen through sandwich and/or competitive immunoassays. The concentrations varied from 10 - 100 ng/ml. The near-infrared dye labeled antigen - antibody complex is excited and the emitted fluorescence is collected with a silicon photodiode detector equipped with an 820-nm bandpass filter. In order to determine various factors influencing the immunosensors performance, the fluorescence intensity responses are obtained under a variety of conditions. The sensor response depending on the type of surface-activating reagent, surface activation period, incubation time, and other measurement conditions also are discussed.