Fluoroimmunoassay

The discovery by Weissman in 1942 that europium (III) complexes of B-diketone O-donor chelates absorb light in the UV region but emit light in the visible region set in train research on the analytical applications of fluorescent f-block complexes that continues to this day. These complexes work by light being absorbed by the ligand (at~360 nm) which undergoes promotion to an excited state from which there is a transfer to metal orbitals that also leaves the metal in an excited state; energy is released at a different wavelength (as visible light at ~615 nm) when the complex reverts to its normal or ground state. The lanthanide ions that can fluoresce strongly are Sm³⁺. Eu³⁺. Tb3+ and Dy³⁺. because the excitation energy level of these four metals lies slightly lower than the excited levels of the ligands and thus they are able to readily accept the energy transfer. With water ligands in particular a mechanism for rapid decay of the excited state other than by fluorescence operates and so it is only in the presence of appropriate ligands that the lifetime of the excited state is sufficient to lead to strong fluorescence, with applicability assisted by a sharp emission peak. The emission process for the complexes is relatively slow and the complex is said to have a long fluorescent lifetime (albeit still in the sub-second range) attributed in part to the 4f orbital being shielded by outer orbitals like the 5s. This long lifetime is a key to analytical applications, since a process is employed that waits a short time following an excitation pulse to let other unwanted but short-lived fluorescence die away, allowing the lanthanide-based fluorescence to be measured essentially free of interference; these are called time-resolved fluorescence analysis methods. Typically, a 'wait time' of ~200 μs followed by a measurement time of ~400 us is employed, permitted because the fluorescent life span of the lanthanide complex is hundreds of microseconds making it possible to detect the target fluorescence with good sensitivity after fading away of the short-lifespan background fluorescence of impurities and organic species.

Fluor immunoassay makes use of the above behaviour. One of the common commercial methods is dissociation-enhanced fluor immunoassay (DELFIA). In this, a nonfluorescent Eu (III) EDTA-like complex is attached by a simple chemical reaction to an antibody or antigen, in a process called labelling. An immunoreaction is next initiated to bind the target, and then a B-diketone and trioctylphosphine oxide (TOPO) mixture are added to the immunocomplex formed, at pH ~3, to promote release of the Eu (III) from the antibody and its complexation as the strongly fluorescent complex [Eu(B-diketonate); (TOPO)2] which is then measured by time-resolved fluorescence methods. The signal size relates to the amount of europium complexed, which in turns relates directly to the amount of the specifically formed target immunocomplex. This process is represented schematically in Figure 9.5. The ease of handling tied to the high detection sensitivity of fluorescent lanthanide probes has led to their being used in base sequence analysis of genes DNA hybridization assay and fluorescent bioimaging apart from in immunoassay. They are rapidly replacing organic

Figure 9.5 Chemistry involved in the fluoroimmunoassay process. Europium ion is bound to an EDTA-like pendant on an antibody, which traps the target pathogen at a second surface-anchored antibody with high specificity. Following washing to remove the test solution, the europium in the trapped adduct is released on chelate addition to form a highly fluorescent complex in solution that is sensed by fluorescence spectroscopy. Signal intensity is directly related to target pathogen concentration.

fluorescent dyes, which have the disadvantage of short fluorescent lifetimes, which means they are compromised by interference signals. This is a fine example of simple coordination compounds at work.

Understanding the photochemistry and photophysics of coordination complexes resulting from the interaction of complexes with light provides a fundamental base for development of other applications. Apart from biomedical imaging and therapy applications of the type exemplified above, applications under active development or the subject of preliminary research include the use of coordination complexes in photocatalysis, optical information transfer and storage, optical computing, analytical sensing and harvesting of solar energy. The penultimate of these fields is exemplified below.

اخر الاخبار

اخبار العتبة العباسية المقدسة

قسم صناعة الشبابيك يواصل أعمال تصنيع بابٍ لحرم مرقد الإمامين الجوادين (عليهما السلام)

قسم العلاقات ينظّم برنامجًا ثقافيًّا لوفد من طلبة مجموعة العميد

قسم الشؤون الفكرية وجامعة العميد يصدران العدد 37 من مجلة تسليم المحكمة

متحف الكفيل يواصل أعمال تركيب عارضات مقتنيات متحف العتبة العسكرية المقدسة

المزيد

الآخبار الصحية

علاج طبيعي لمشكلات البشرة وحب الشباب

اكتشاف ارتباط عنصر شائع في نظامنا الغذائي بحصوات المرارة

كل ليلة.. 11 دقيقة إضافية من النوم لها مفعول السحر

علاج طبيعي يخفف أعراض سكري الحمل

المزيد

الآخبار التكنلوجية

قمر وردي ومذنب يلامس الشمس.. 8 ظواهر فلكية تزين السماء طوال شهر أبريل

تفسير حديث لتحيز التوحد بين الجنسين.. هل تمتلك الإناث "درعا جينيا" ضد المرض؟

الجلوس الذهني النشط.. مفتاح تقليل خطر الخرف لدى كبار السن

خسائر غير مسبوقة للجليد في أيسلندا وسواحل المحيط الهادئ خلال 2025

المزيد

مواضيع ذات صلة

Fluoroionophores

How Much? - Analysing with Complexes

Other Metallodrugs

Anticancer Drugs

A Laboratory-free Approach - In Silico Prediction

Doing What Comes Unnaturally– Synthetic Biomolecules

Mixed-Metal Proteins

Other Metal-containing Biomolecules

Zinc-containing Biomolecules

Copper-containing Biomolecules

Iron-containing Biomolecules

Metalloproteins and Metalloenzymes