Rapid quantitative assay for erythrocyte porphyrins

Manganese superoxide dismutase depletion in murine hematopoietic stem cells perturbs iron homeostasis, globin switching, and epigenetic control in erythrocyte precursorcells

“…Free macrocycle assays were performed as previously described [30]. Briefly, equal volume or mass of tissue were combined with Celite (Sigma Aldrich, St. Louis, MO) to remove particulate matter.…”

Distribution of non‐heme porphyrin content of individual erythrocytes by fluorescence image cytometry and its application to lead poisoning

“…In the present work a 1 mm thick fluorescent glass slide (Fish-Schurman Corp., New Rochelle, NY) was used as a secondary fluorescence intensity standard, to correct for day-to-day variations in the excitation light intensity, By focussing on its lower surface its fluorescence intensity, Ist, was measured in the course of each experiment; all data were then normalized by dividing all cytometric intensities by I,, and were expressed in fluorescence units (f.u.). A calibration of the total fluorescence intensity emitted by an individual cell, Irbcr in terms of its ZPP content was obtained by relating the average fluorescence derived from a specimen's distribution function, I,,, to the ZPP level of the same specimen, determined either fluorometrically or by an acid extraction assay (17).…”

“…The quantitation of non-heme porphyrin has long been recognized as a n important diagnostic tool in these conditions (9) and the level of zinc protoporphyrin (ZPPj in blood is, for example, widely used in screening populations a t risk for undue lead exposure, such as various occupational groups and young children. In these porphyrin-based assays the average concentration of non-heme porphyrin in the erythrocytes is determined, either by solvent extraction (17) or, more usually, by a fluorometric method (lo), including the use of dedicated hematofluorometers (4,5). These fluorometric assays are based on the fact that free (unchelated) erythrocyte porphyrin (FEP), as well as Zn2 ' -chelated porphyrins (ZPP), fluoresce with significant quantum yields in the 600 nm region, while the heme is practically non-fluorescent, since the iron ion effectively quenches the red porphyrin fluorescence (7,151. The present paper describes an image-based methodology for measuring the distribution of the non-heme porphyrin content of individual erythrocytes and addresses the question of what information about the underlying pathological condition may be obtained from it.…”

Bestimmung der Erythrozyten-Porphyrine

“…An die Stelle der älteren Extraktionsmethode, die recht zeitaufwendig ist und 10-15 ml Vollblut erfordert, traten in den letzten Jahren Mikromethoden, die in kürzerer Zeit durchführbar sind und nur 20 111 Vollblut erfordern (1,3,5,6).…”

“…In einer Stunde lassen sich ohne Schwierigkeiten 30 Bestimmungen durchführen und mit einem Drucker dokumentieren. Das Hematofluorometer ZnP 400 ist aus diesen Gründen besonders für Screening-Untersuchungen an thode nur Abweichungen von 2,5% (5). Die Wiederfindung bei der oben beschriebenen Methode beträgt 95%, bei der Wranne-Methode nur etwa 80%.…”