Effect Of Storage Time And Temperature On Cell Morphology, Quantity, And Degeneration Rate Of Non-Fixed Pleural Effusion And Ascites

Objective:Along with the wide application of pleural effusion and ascites cytology diagnosis,how to reduce missed diagnosis,misdiagnosis,improve the diagnostic positive rate has important practical significance.Saved correctly,timely submission and reasonable fixation is an important premiseis for assurance cytologic diagnosis correctness.This study intends to analysis the cell morphology,cell degeneration of the mononuclear cells,mesothelial cells and tumor by the conventional cytology smear and cell blocks in different storage environment.We evaluated numerical and morphological changes in pleural fluid specimens processed after storage at room temperature or under refrigeration to provide theoretical basis for the preservation environment of hydrothorax and ascites specimens and the best time to produce cell smears and cell blocks.Method:We collected 30 cases of hydrothorax and ascites specimens from The Second Hospital Of Hebei Medical University Department of pathology in 2015-2016.Each fresh specimens was fully mixed and divided evenly into 18 copies,each 10 ml,sealed centrifuge tube.Two specimens were immediately centrifuged and made cell smears and cell blocks respectively,and recorded as control group.The remaining 16 specimens were divided into the following groups: 8 samples were refrigerated environment group(4℃),8 samples were room temperature environment groups(24℃).In each case,half of these tubes(eight tubes)were maintained at room temperature;the other eight tubes were refrigerated at 4 ℃.The samples were processed after 6 hours and after 6 hours,24 hours,48 hours and 72 hours from thoracocentesis.For all samples the following procedures were adopted.The fluid was then centrifuged at 3000 rpm for 10 minutes.After supernatant removal,the samples were smear slides and the sediment cells made into cell blocks.The slides were stained with a haematological stain.Results:1 Through qualitative observation of cell morphology in cell smears and cell blocks,we found that the cell structure was clear,the size and shape of the cells was normal in the control group,and the cell types can be distinguished.With the extension of time,cells occurred a series of degenerative changes.Morphological cell changes appeared such as cell volume increased,nuclear staining faded,structure of chromatin changed,or cell volume decreased,nuclear pyknosis and cytoplasmatic alteration,it was difficult to distinguish the cell morphology.The classical tubular gland or mesh structure gradually disappeared and became unrecognized degenerative cell masses in parafin sections of cell blocks.Compared to room temperature group,the cell morphology have a slight advantage in the refrigeration environment group at the same time.2 Observation results showed that the average number per high power field of cell smears was gradually reduced in 72 hours both in room temperature(r=-0.321,P<0.05)and in refrigerated conditions(r=-0.357,P<0.05)with the extension of time in vitro.In the temporal analysis,for both temperatures,the average number per high power field of cell smears was significantly decreased at 48 hours and 72 hours when compared with the control group,but there was no significant difference at 6 hours and 24 hours.The statistical results had no significant difference between the refrigerated conditions and the room temperature conditions at the same time.3 With the extension of time in vitro,the degeneration rate of cells in hydrothorax and ascites significantly increased in 72 hours both in room temperature(r=0.850,P<0.05)and in refrigerated conditions(r=0.654,P<0.05).In the temporal analysis,for both temperatures,the degeneration rate of cells in hydrothorax and ascites significantly increased at 6 hours,24 hours,48 hours and 72 hours when compared with the control group.In 6 hours groups,the degeneration rate of cells in refrigerated conditions significantly lower than in room temperature,the results have statistical significance.Compared with the groups at the same 24 hours、48 hours and 72 hours,the statistical results had no significant difference in refrigerated conditions and in room temperature.4 During the experimental period,bacterial contamination occurred in 4 samples of 30 cases with pleural effusion and ascites,13.33% of the total number of samples.In room temperature environment,two cases of bacterial contamination appeared at 24 hours,one at 48 hours and one at 72 hours.In refrigerated conditions,one case of bacterial contamination appeared at 24 hours,one at 48 hours and two at 72 hours.Conclusion:1 Observation results showed that the average number per high power field of cell smears was gradually reduced and the degeneration rate of cells in hydrothorax and ascites significantly increased in 72 hours both in room temperature and in refrigerated conditions with the extension of time in vitro.2 There was no significant difference at 6 hours in the average number per high power field of cell smears between the refrigerated conditions and the room temperature conditions.In 6 hours groups,the degeneration rate of cells in refrigerated conditions was significantly lower than in room temperature.It is more suitable for preservation of non-fixed pleural and ascitic fluid samples in the refrigeration environment than in the room temperature environment.