Assaying Granulocyte Phagocytosis by Chemiluminescence: Effect of Storage Time and Temperature of Blood Samples

Abstract

The objective was to study the effect of storage of blood samples for up to 4.5 h at either room temperature or on ice on granulocyte phagocytosis as assessed by cheduminscence. The study included three groups of animals: eight pigs, seven cows and eight horses, which were bled at one occasion. The blood samples were divided into two and stored either at room temperature or on ice. Granulocyte phagocytosis in the samples was then assessed every 60 min starting 30 min and finishing 4.5 h after bleeding. The phagocytosis was recorded as the AUC of chemiluminescence following phagocytosis of opsonized zymosan particles. The time of storage of blood samples had a significant (p < 0.001) overall effect on the chemiluminiscence following phagocytosis in all species tested, as was the case with storage temperature (P < 0.001). It should be noted, however, that there were considerable species differences in the effect of the storage on the granulocyte phagocytosis. Hence, for valid comparisons of data on granulocyte phagocytosis assessed by chemiluminiscence it is important that the handling procedures of the blood samples have been applied consistently even if the assays have been performed only a few hours after blood collection.

The effect of storage on whole blood chemiluminescence measurement of equine neutrophils

“…It has been shown that isolated human neutrophils exhibit lower light emission as early as 2 h after storing at room temperature, and after 24 h, it is almost nonexistent (17). Conversely, cells stored for 4 h at 4 C showed the same light emission as fresh ones and they retained one third of their initial activity after 24 h. Similar results have been obtained in whole human blood (18) and in samples collected from horses, cattle and pigs (19), demonstrating that to guarantee optimal measurement results, blood should be stored at 4 C and assays should be performed no later that 3, 4, 1 and 1 h(s) after collection, respectively.…”

“…It is generally accepted that CL values in whole blood reflect homeostatic changes in the organism better than studies on isolated leukocytes, and thus are more useful in clinical tests . Moreover, whole blood assays do not require laboratory procedures to isolate neutrophils, which may influence survival rate, activity and expression of receptors in these cells .…”

“…However, findings are not always applied to animal studies. This has been shown by the wide influence of storage duration on CL values in blood of different breeding animal species as well as wild animals . The aim of the current study was to determine the effect of duration and temperature of sample storage on whole blood CL measurement results.…”

Induced Parturition in the Cow – a Suitable Model for Studies of Postpartum Disturbances

“…In PG‐induced cows uterine ultrasonography was not performed. The phagocytosis capacity was assessed by measuring the chemiluminescence following phagocytosis of opsonized zymosan particles and the spontaneous respiratory burst of the PMN was assessed by recording the chemiluminescence in samples without zymosan according to Magnusson and Holst (1998). Using a luminometer (ML 3000, Dynatech Laboratories, In Vitro, Solna, Sweden).…”

Oxidant/Antioxidant Status Assessment of Blood in Selected Equine Diseases

“…Chemiluminescence in whole blood was analysed on the basis of the assumptions of Magnusson and Holst (18) stating that measurements in whole blood are a better reflection of homeostasis in vivo than examinations performed on isolated leukocytes and are thus more useful in clinical trials. Measurements in whole blood also do not require any laboratory procedures to isolate neutrophils, which could affect their survival, or expression and activity of cell surface receptors (23).…”