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Comparison of devices and microscopical assesment differential count of leukocytes
NETREFOVÁ, Veronika
This thesis deals with methods of determination of differential count of leukocytes and their positives and negatives. No wonder that nowadays modern haematology analyzers have been used for determination of differential count of leukocytes due to technical progress, however manual assesment by microscope keep still being a basic element of good work in a haematology laboratory. In the practical part I will describe single work procedures and these methods will be then compared according to the observed results. The Coulter LH 750 analyzer by Beckman Coulter Inc. based on the principle of electric impedance has been used in Nemocnice České Budějovice a.s. for automatic assesment of differential count of leukocytes. In case of the microscopic assesment a blood film is made which is then stained by Pappenheim method with May - Grünwald and Giemsa - Romanovsky colours. Then I will try to suggest an optimization for current investigation of the complete blood picture.
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Circadian changes of Spodoptera littoralis haemocytes
TREFILOVÁ, Terezie
Abstrakt This work is a study of the morphology of hemocytes and stability of the basic non-pathological hematological levels during 24 hours. Spodoptera littoralis. (Boisd.) (Lepidoptera: Noctuidae) in the sixth instar was used as the biological model. Light microscopy was the technique used to observe the hemocytes. Prior to the microscopy, preparations were made from the hemolymph and stained by Pappenheim's staining method. This is a method which is routinely used in veterinary as well as human laboratory practice in hematology. Six types of hemocytes were observed: small granulocytes (sGR), large granulocytes (lGR), small prohemocytes (sPR), large prohemocytes (lPR), spherulocytes (SP), and plasmatocytes (PL). Pappenheim's staining also enabled apoptosis to be followed. The knowledge of circadian changes in hemocytes in S. littoralis is very important if this animal is to be employed as a new biological model, for instance, for investigations in imunology, hematotoxicology or pharmacotoxicology. Changes in the levels of some substances can be physiological even if lying beyond the reference level limits (Knapp et al., 1984). We found the various hemocyte counts varying throughout the day. At midnight, the sPR and lPR counts are lowest while the apoptotic cell and lGR counts are highest. We tested the use of an ELISA reader (Sunrise, Tecan) (which is based on the photometric principle) with a view to increasing the precision and shortening the time of analysis. However, due to the large spans of concentrations, viz. roughly from 2 mmol/l to 30 mmol/l, the use of the ELISA reader was found unsuitable.
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Morfocytometric investigation of haemocytes in comparative hematology
ŠUBJAK, David
Standard haematological procedures in preclinical toxicity studies are mostly carried out on mammals, more recent haematotoxicological models represent invertebrates because they can be used in the screening of myelotoxicity. In the present study, we examined haemocytes of Culex pipiens as a new potential alternative haematological in vivo model. Preparations for microscopic analysis were made from the hemolymph stained by Pappenheim's method for overstriking morphology and by toluidin blue and AgNOR for overstriking nucleoles. Microscopic photos were processed by image analysis and size of cells, nuclei and nucleoli were evaluated. Six types of haemocytes were found: granulocytes, prohemocytes, spherulocytes, plasmatocytes, adipohemocytes and oenocytes. Thanks to image analysis we made a morfometric profile of each type of haemocytes and set the nucleolar coefficient. We described significant differences among prohemocytes, plasmocytes and oenocytes size. On the grounds of nucleoles size we divided haemocytes to next two significant groups. Image analysis seems to be very helpful for distinction among various haemocyte lines but we cant use it for generally separation without personal control thanks to presence of statistically significant findings for only some comparisons cell types.
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Examination of changes in fagocytosis for the screening of hemotoxicity on a biomodel
JURČOVÁ, Martina
This thesis dealt with the study of laboratory methods used in the examination of phagocytosis and we conducted the research of the examination of the changes of phagocytosis on the biomodel S. littoralis. The results showed the assumed increase in phagocytosis after the administration of the model substance. Our tests applied documented quite sensitively the changes in fagocytic acitivity and production of cytotoxic substances and the model animal was evaluated as a suitable biomodel for the other studies of hemotoxicity.
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