Comparison of manual and automated cell counts in EDTA preserved synovial fluids. Storage has little influence on the results

Abstract

Objective-To determine the precision and agreement of synovial fluid (SF) cell counts done manually and with automated counters, and to determine the degree of variability of the counts in SF samples, kept in the tubes used for routine white blood cell (WBC) counts-which use liquid EDTA as anticoagulant-at 24 and 48 hours at 4°C, and at room temperature. Methods-To determine precision, cell counts were repeated 10 times-both manually and by an automated counter-in a SF sample of low, medium, and high cellularity. The variances were calculated to determine the interobserver variation in two manual (M1,M2) and two automated cell counts (C1,C2). The agreement between a manual (M1) and automated counter (C1) results, was analysed by the Bland and Altman method and the diVerence against the mean of the two methods was plotted. Then, the mean diVerence between the two methods was estimated and the standard deviation of the diVerence. To determine the eVects of storage, SF samples were kept in a refrigerator at 4°C, and at room temperature; cell counts were done manually (M1) and automatically (C1) at 24 and 48 hours and the changes analysed by the Bland and Altman method. The variances were compared using an F test. Results-(1) Precision. With the manual technique, the coeYcients of variation were 27.9%, 14%, and 10.7% when used for counting the SF with low (270), medium (6200), and high cellularities (25 000). With the automated technique the coeYcients of variation were 20%, 3.4%, and 2.9% in the same SF samples. In the fluids of medium and high cellularity, the variances of the automated cell counts were significatively lower (F test, p<0.002) than those of the manual counts. (Ann Rheum Dis 1997;56:622-626) The white blood cell (WBC) count in synovial fluid provides information about the amount of inflammation present in the joints, and is the basis for the classification of synovial fluids (SF) into the categories of non-inflammatory, inflammatory, and septic.1-3 The WBC count is generally performed manually, using saline as a diluent to avoid the formation of a mucin clot, which would result from the use of haematological diluents containing acetic acid. 4 DiVerent problems have been found in relation to the performance of the WBC counts in the SF; a poor reproducibility of manual WBC counts, 5 as well as contradictory results in relation to the stability of the SF have been reported, emphasising the need of performing the procedure without delay. 6 Counting by means of automatic counters has occasionally been done, but problems such as the error produced by fat globules have been found when the counts are performed by such a method. 7Our aim is to determine the precision and reproducibility of WBC counts of SF

The added value of synovial fluid centrifugation for monosodium urate and calcium pyrophosphate crystal detection

“…Synovial fluids were maintained for at least 48 h without morphological change. Even after one to several weeks of storage at 4°C, the crystal and leukocyte concentrations only tend to be minimally reduced [12,[14][15][16][20][21][22]. Therefore, we think that our storage handling had none or minimal influence on crystal recognition.…”

“…In vitro studies suggest that a reduction of even 2°C is sufficient to lower the solubility point [19]. Although, our preservation method, in a refrigerator at 4°C, is supported by a handful of small studies which assessed the impact of storage handling on crystals recognition [6,[12][13][14][15][16][17]. Synovial fluids were maintained for at least 48 h without morphological change.…”

“…Both tubes, filled with 1 ml of SF, were stored in a refrigerator at 4°C awaiting microscopy [6,[12][13][14][15][16][17]. SF smears were microscopically examined before and after centrifugation, as soon as possible, within 5 days as target:…”

Dried cytospin preparations of synovial fluid are a stable material for long-time storage and delayed crystal analysis

“…For scientific investigations, in controversies as well as the distribution of SF to other laboratories for secondary opinion, the possibility of storage and delayed examination for crystals may be beneficial [2]. Several studies assessed the impact of storage conditions and duration on the number and morphology of monosodium urate (MSU) and calcium pyrophosphate dihydrate (CPPD) crystals [1,[3][4][5]. Crystals can be maintained for prolonged periods without morphological changes by freezing [3,5], which requires refrigerators with a certain storage capacity and is not practicable for sample distribution.…”

Review Article: Joint Aspiration for Diagnosis of Periprosthetic Infection

“…29 Synovial fluid should be stored in tubes containing ethylenediaminetetraacetic acid (EDTA) and kept the specimen at 4ºC before analysis. 30 Synovial fluid white cell count decreases by 47% after 48 hours when stored in heparin, compared with 5.1% when stored in EDTA. EDTA is a much more suitable preservative than heparin.…”