EVALUATION OF THE DELTA INSTRUMENTS SOMASCOPE SOMATIC CELLS COUNTER

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1 EVALUATION OF THE DELTA INSTRUMENTS SOMASCOPE COUNTER The SomaScope LFC is a automatic istumet, which pemits the eumeatio of somatic cells i milk. It is maufactued by Delta Istumets (Advaced goup, Nethelads) ad distibuted by Humeau i Face. This istumet uses the fluoo-opto-electoic method. The test potio is mixed to a dye (DAPI) i ode to dispese fat globules ad to dye somatic cell uclei. A aliquot is ijected ito a lamia flow caie fluid. The cells staied ae sepaated by the flow ad ae exposed to the light beam of a Light Emittig Diode (LED) ad emit a yellow fluoescece light diected towad 2 detectos. Oly impulses, whose the itesities ae beyod to a fixed theshold (i mv) ad eceived by both detectos, ae couted ad coveted i tem of cellula cocetatio usig a calibatio equatio. This appaatus, which is coected to a compute that esues the sigal teatmet, ca be associated to the "Lactoscope" ifaed aalyse to costitute the "Combiscope". The evaluatio tests wee pefomed i ACTALIA Cecalait's physico-chemisty laboatoy (efeece aalyses ad implemetatio of the SomaScope) fom Febuay to Jue 213. Afte pelimiay tests of stability of the istumet, cotamiatio betwee samples, lieaity ad calibatio, the epeatability ad accuacy wee evaluated i cow, goat ad ewe milks. A cleaig solutio (aqueous solutio of Deco at 4 %), a caie fluid (aqueous solutio of tito X1 at,1 %), ad a dye solutio (ef S7 kit) wee ecessay fo these tests. The appaatus was cofigued fo a ate of 6 samples pe hou ad o coectio of cotamiatio. The appeciatio citeia of the estimated paametes wee take fom ISO / IDF 128-3:21, o fom the CNIEL/IE hadbooks coceig the use of somatic cells coutes with the cotext of milk paymet ad milk cotol i Face (CNIEL PROC CE 4). A. PRELIMINARY TESTS A.1. Evaluatio of the stability The stability was evaluated by the aalysis, i automatic mode, of milk evey 2 miutes, epesetig 2 measuemet cycles. This evaluatio was pefomed o 3 levels of cellula ates 3 times pe level. To evaluate the stability of the istumet, the epeatability ad epoducibility wee calculated fo each level.. The stadad deviatio of epoducibility values (3.1 ; 2.2 ad 2.1) ae i accodace with the ISO / IDF stadad (SR 5 %).

2 A.2. Evaluatio of cotamiatio betwee samples This citeio was evaluated i automatic aalysis mode, by aalysig the same cow milk ad distilled wate accodig to the sequece: MILK - MILK WATER WATER epeated twety times o 3 levels of cellula ates The cotamiatio ates betwee successive samples ae i accodace with the ecommedatios of the ISO / IDF ad the CNIEL PROC CE 4 hadbook (Tc 2 %). A.3. Evaluatio of lieaity The samples wee pepaed by mixig the poteic etetate ad filtate obtaied by tagatial ultafiltatio (cutoff theshold:,8µm). A age of 2 levels, egulaly distibuted fom to 25.1 E 3 cells/ml, was pefomed by weighig ad the coespodig dilutios wee calculated usig the covesio of the desities. Each level was aalysed 5 times i maual mode. The A/At atio (A ad At: amplitude of esidues ad amplitude of cotet espectively) equal to 2.54 % is highe tha the ecommedatios (2 %) of the ISO / IDF stadad. This value ca be explaied by a dispesio of the esults athe tha a lieaity defect. A F test, which compae the liea ad ode 2 cuviliea egessio, cofims this aalysis (Fobs = 1.7 < F (.95,18,63) = 1.77 at 5 % isk) The lieaity of the istumet is satisfactoy fo the E 3 cells/ml age. A.3. Evaluatio of the calibatio The evaluatio of the calibatio, iitially istalled by the maufactue, was pefomed with 1 commecial somatic cells efeece mateials (SRMs) poduced by ACTALIA Cecalait i Jue 213. Each sample was aalysed i duplicate. The esults peset a low mea ad stadad deviatio of deviatios. The esidual stadad deviatio is equivalet to the stadad deviatio of deviatios. The mea bias (1.2 %) ad the egessio slope (1.5) ae i accodace with the ecommedatios of the ISO / IDF stadad ad the CNIEL PROC CE 4 hadbook (espectively +/-5 % ad 1 +/-.5). Moeove the esidual stadad deviatios of liea egessio obtaied ( E 3/ml) is i accodace with the ecommedatios of the CE 4 (Sy,x 2.1 E 3/ml). B. EVALUATION OF REPEATABILITY AND ACCURACY B.1. The samples The tests wee pefomed usig: fo cow milk: 8 samples of of hed milk fom the Fache-Comté egio ad 12 samples of idividual milk fom 4 fams i the Jua. Boopol was added to the idividual milk samples to give a fial cocetatio of.2 %. fo goat milk: 1 samples of hed milk fom the Poitou-Chaetes egio. fo ewe milk: 8 samples of hed milk fom the Roquefot su Soulzo egio. B.2. Pocedue The epeatability ad accuacy of the istumet wee evaluated usig all the samples (hed ad idividual milk samples). The quatitative aalyses wee pefomed i automatic aalysis mode, i duplicate fo each set of 2 samples accodig to the followig sequece: Set 1 ep 1 - Set 1 ep 2 - Set 2 ep 1 - Set 2 ep 2 Set ep 1 - Set ep 2. A cotol milk was aalysed evey 2 samples to veify the stability of the aalyse. The accuacy evaluatio coces the values obtaied afte calibatio ad adjustmet of the istumet with commecial SRMs poduced by ACTALIA Cecalait. The efeece method : 27 (fo cow ad ewe milk, ad accodig to the ifomative aex fo goat milk) was used fo the eumeatio of the somatic cells by micoscope coutig (simple measuemet cofimed if impotat esidue). B.3. Results

3 B.3.1. Cow milk The followig tables ad figues peset the esults obtaied: Somatic cells age Mi Max M Sx S S (%) Global If Table 1: SomaScope epeatability citeia i cow hed milk samples Somatic cells age Mi Max M Sx S S (%) Global If Table 2: SomaScope epeatability citeia i cow idividual milk samples : umbe of esults; mi ad max: miimum ad maximum values; M ad Sx: mea ad stadad deviatio of the esults; S ad S %: absolute a elative stadad deviatio of epeatability; : maximum deviatio of epeatability o 95 % of cases it ca be oted that: Fo hed milk: the global elative stadad deviatio of epeatability (3.77 %) is i cofomity with the ecommedatios of the ISO / IDF stadad (S 4 %) ad of the CNIEL PROC CE 4 hadbook (S 6 %). Fo the media age (11-.1 E 3/ml), coespodig to the majoity of the esults, the elative stadad deviatio of epeatability obtaied (3.66 %) is also i accodace with the ecommedatios of ISO / IDF stadad (S 4 %). Fo idividual milk: the acceptability limits ae the same tha fo hed milk. The global elative stadad deviatio of epeatablity (7.19 %) is highe tha the limits due to a lot of vey low esults. Howeve, fo the media age (11-.1 E 3/ml), which epesets the most cuetly esults obseved, the elative stadad deviatio of epeatability (3.3 %) is i cofomity. Somatic cells Hed milk Somatic cells Idividual milk mi max Y Sy d Sd Sy,x Sy,x (%) Table 3: SomaScope accuacy i cow milk sample, mi, max: umbe of esults, miimum ad maximum values; Y: mea esults usig the efeece method; Sy: stadad deviatio of the esults fom the efeece method; d, Sd: mea ad stadad deviatio of deviatios; Sy,x: esidual stadad deviatio of the liea egessio (istumet vs efeece).

4 12 COW HERD MILK 8 6 y =,987x - 8,8 4 2, 2, 4, 6, 8,, 12, SOMASCOPE 25 2 COW HERD MILK 15 y = 1,6x + 2, SOMASCOPE Figues 1 et 2: Relatio betwee SomaScope ad efeece esults i cow milk It ca be oted that: Fo hed milk: the mea ad stadad deviatio of deviatios ae espectively equal to 12.7 ad E 3 cells/ml. The egessio slope (.987) ad the itecept (-8.8) obtaied ae ot sigificatly diffeet fom espectively 1. ad zeo (P = 5 %). The esidual stadad deviatio of egessio ( E 3 cells/ml = 8,11 %) is i accodace with the ecommedatios of the ISO / IDF stadad (Sy,x 1 %). Fo idividual milk: the mea ad the stadad deviatio of deviatios ae espectively equal to 3.3 ad E 3 cells/ml. The egessio slope (1.6) ad the itecept (2.2) obtaied ae ot sigificatly diffeet fom espectively 1. ad zeo (P = 5 %). The esidual stadad deviatio of egessio ( E 3 cells/ml = 8,45 %) is i accodace with the ecommedatios of the ISO / IDF stadad (Sy,x 1 %). Fo cow milk, the esults obtaied ae i accodace with the ecommedatios of the ISO / IDF stadad. B.3.2. Goat milk The followig tables ad figues peset the esults obtaied: Somatic cells age Mi Max M Sx S S (%) Global If Table 4: SomaScope epeatability citeia i goat milk samples

5 : umbe of esults; mi ad max: miimum ad maximum values; M ad Sx: mea ad stadad deviatio of the esults; S ad S %: absolute a elative stadad deviatio of epeatability; : maximum deviatio of epeatability o 95 % of cases The global elative stadad deviatio of epeatability (2.82 %) is i accodace with the ecommedatios of the ISO / IDF stadad (S 4 %) ad the CNIEL PROC CE 4 hadbook (S 3 %). Fo the high age ( E 3/ml), coespodig to the majoity of the esults, the elative stadad deviatio of epeatability (2.61 %) is slightly highe tha the ecommedatios of the ISO / IDF stadad (S 2 %). mi max Y Sy d Sd Sy,x Somatic cells Table 5: SomaScope accuacy i goat milk sample, mi, max: umbe of esults, miimum ad maximum values; Y: mea esults usig the efeece method; Sy: stadad deviatio of the esults fom the efeece method; d, Sd: mea ad stadad deviatio of deviatios; Sy,x: esidual stadad deviatio of the liea egessio (istumet vs efeece). Sy,x (%) 3 25 GOAT MILK 2 15 y =,999x - 22,72 5, 5,, 15, 2, 25, 3, SOMASCOPE Figue 3: Relatio SomaScope ad efeece esults i goat milk samples It ca be oted that the mea ad the stadad deviatio of deviatios ae espectively equal to 22.8 ad E 3 cells/ml. The egessio slope (.999) ad the itecept (-22.2) obtaied ae ot sigificatly diffeet fom espectively 1. ad zeo (P = 5 %). The esidual stadad deviatio of egessio ( E 3 cells/ml = 3,7 %) is i accodace with the ecommedatios of the ISO / IDF stadad (Sy,x 1 %). Fo goat milk, the esults obtaied ae i accodace with the ecommedatios of the ISO / IDF stadad. B.3.3. Ewe milk The followig tables ad figues peset the esults obtaied: Somatic cells age Mi Max M Sx S S (%) Global If Table 6: SomaScope epeatability citeia i ewe milk samples : umbe of esults; mi ad max: miimum ad maximum values; M ad Sx: mea ad stadad deviatio of the esults; S ad S %: absolute a elative stadad deviatio of epeatability; : maximum deviatio of epeatability o 95 % of cases The global elative stadad deviatio of epeatability (2.71 %) is i accodace with the ecommedatios of the ISO / IDF stadad (S 4 %) ad the CNIEL PROC CE 4 hadbook (S 4 %). Fo the media ag (11-.1 E 3/ml), coespodig to the majoity of the esults, the elative stadad deviatio of

6 epeatability obtaied (3.2 %) is also i accodace with the ecommedatios of the ISO / IDF stadad (S 4). mi max Y Sy d Sd Sy,x Somatic cells Table 7: SomaScope accuacy i ewe milk sample, mi, max: umbe of esults, miimum ad maximum values; Y: mea esults usig the efeece method; Sy: stadad deviatio of the esults fom the efeece method; d, Sd: mea ad stadad deviatio of deviatios; Sy,x: esidual stadad deviatio of the liea egessio (istumet vs efeece). Sy,x (%) EWE MILK y =,953x + 4,81 4 2, 2, 4, 6, 8,, 12, 14, 16, SOMASCOPE Figue 4 : Relatio betwee SomaScope ad efeece esults i ewe milk The mea ad the stadad deviatio of deviatios ae espectively equal to 14.1 ad E 3 cells/ml. The egessio slope (.953) is sigificatly diffeet fom 1. (P = 1 %) ad the itecept (4.8) is ot sigificatly diffeet fom zeo (P = 5 %). The esidual stadad deviatio of egessio ( E 3 cells/ml = 4,55 %) is i accodace with the ecommedatios of the ISO / IDF stadad (Sy,x % 1 %). Fo ewe milk, the esults obtaied ae i accodace with the ecommedatios of the ISO / IDF stadad. CONCLUSION The esults obtaied duig this evaluatio ae i accodace with the ecommedatios of the ISO / IDF 128-3: 21 stadad ad/o the CNIEL/IE hadbook coceig the use of the somatic cells coute withi the cotext of milk paymet ad milk cotol i Face (CNIEL PROC CE 4) fo the 3 types of milk (cow, goat ad ewe). Accodig to the evaluatio epot of the Delta Itumets SomaScope somatic cells coute X. QUERVEL, P. TROSSAT Septembe 213