COUNTING OF SOMATIC-CELLS IN MILK - STUDI ES ON THE MEASUREMENT USINGFLOW-CYTOMETRY (SOMACOUNT) AND COMPARISON WITH THE MEASURING RESULTS OBTAINED BY MEANS OF THE FLUORESCENCE OPTICAL-SYSTEM (FOSSOMATIC-360)
Wh. Heeschen et al., COUNTING OF SOMATIC-CELLS IN MILK - STUDI ES ON THE MEASUREMENT USINGFLOW-CYTOMETRY (SOMACOUNT) AND COMPARISON WITH THE MEASURING RESULTS OBTAINED BY MEANS OF THE FLUORESCENCE OPTICAL-SYSTEM (FOSSOMATIC-360), Kieler Milchwirtschaftliche Forschungsberichte, 46(4), 1994, pp. 291-316
For counting somatic cells in milk (''cell counting'') as part of the
EC/EU- and national regulations standardization is indispensable; this
includes, in particular, the comparability of methods. In the present
study flow cytometry (Somacount) is compared with the fluorescence op
tical system (Fossomatic 360), as well as with optical count (Breed) a
s reference method, the aim being to establish, to what extent the new
method yields comparable results. Precision was calculated from multi
ple counts. In the range below 500 (in 1000 cells/ml) the s.d. of repe
ated countings does not exceed 10, the variation coefficient in the ra
nge between 250 and 1.000 is 2 %, and beyond this range 1.6 %. After p
retreatment using the preservatives potassium dichromate and Bronopol
the samples keep for more than 48 h. With a limitation in time the sam
e results in terms of shelf life are obtained when sodium azide and bo
ric acid are used. The shelf life of the ''raw milk method'' (cold sto
rage at 6-degrees-C) does not lead to temporal stability. The measurin
g results between preservation methods are comaparable, when potassium
dichromate, boric acid or Bronopol were used. With the ''raw milk met
hod'' slightly lower values are obtained. Preservation with sodium azi
de leads to markedly lower values. This is in agreement with the exper
ience gained with other instruments. In the normal measuring range the
carry over error of the apparatus from sample to sample is 1-2 %, and
, hence, of no importance. No problems were encountered with the tempo
ral stability of the measuring method for 4 days using potassium dichr
omate. With plus/minus 2-3 % relative differences (means of repeated c
ountings) are within the range of general accuracy. Using 407 quarter
foremilk samples Somacount and Fossomatic were compared; with untransf
ormed scale a correlation coefficient of 0.998 was found and after log
transformation 0.917. In units of the log scale the residual s.d. is
0.06 in the range between 40 (in 1000 cells/ml) and 2.500, which corre
sponds to a factor of 1.14. This means, the relative error is 14 %. As
to the counting level a 1:1 agreement is found. The optical method's
missing accuracy is disadvantageous for comparisons. Considered all in
all, however, there are no substantial differences in the counting le
vel compared with the Somacount. A first international collaborative t
rial involving 17 laboratories with altogether 26 sections was perform
ed according to the IDF standard 148:1991. In this intercomparison tri
al 10 milks samples in the range between 129 and 1.086 (in 1000 cells/
ml) were used. A first evaluation showed a satisfactory repeatability
(r = 26-103). Reproducibility was poor (R = 62-541) for the 10 milks (
= cell levels). This was, however, solely due to the inexperience of o
ne laboratory only. After exclusion of this laboratory good intercompa
rison test results were obtained (r = 26-95, R = 14-89). The IDF-targe
ts for cv(r) and cv(R) are clearly observed. With a cell level of 428
cv(r) is 4.1 % (target = 4-5 %) and cv(R) 8.1% (target = 10-12%). Henc
e, the Somacount instrument itself and the participating laboratories
fulfil, after adequate training, the targets of the IDF standard 148:1
998. Good Laboratory Practice (calibration of apparatus) is supported
by reference materials, which are treated using thermo-chemical method
s and keep for several months (''Kiel standards''). Microscopically co
ntrolled numbers of cells are available in biological matrix for contr
olling the laboratory equipment. There exist no systematic differences
between the values counted in the standard samples prior to (raw milk
) and after treatment.