[Determination of the number of somatic cells in milk using the rapid diphenylamine DNA filter method]. / Stanovení poctu somatických bunek v mléce rychlou difenylamin DNA filtr metodou.

The rapid reaction of the diphenylamine agent with DNA was used for the determination of the counts of somatic cells in cow's milk, using the DNA filter method. The method is based on the filtration of a warmed (65-70 degrees C) mixture of milk with Triton X-100 through the Synpor nitrocellulose membrane filter, pore size 2 to 5 microns, and subsequent DNA determination of the collected somatic cells by the colour reaction of diphenylamine. A 2ml quantity of distilled water and 4 ml of diphenylamine reagent were added to the membrane filters with somatic cells. The mixture is warmed in water bath at 90 to 100 degrees C for 20 min., then it is cooled, centrifuged (3500 X g, 15 min.), and the optical density is measured at 595 nm. The relation 8 micrograms = 1 million cells was used for the conversion of DNA content to the counts of cells. The average variation coefficient of the determination was 5.9% and the coefficient of correlation between the diphenylamine DNA filter method and the direct microscopy of the somatic cells on membrane filters was r = 0.997. Using the diphenylamine DNA filter method, the counts of somatic cells can also be determined from milk samples stored in frozen condition or from the filters with collected cells kept at the temperature of 4 degrees C (10 days) or 25 degrees C (3 days). Milk stabilized with formaldehyde can also be used for the determination if stored at 4 degrees C.

[Use of the Synpor membrane filter for the separation and determination of the number of somatic cells in the milk of dairy cows using the indole DNA filtration method]. / Vyuzití membránových filtru Synpor pro separaci a stanovení poctu somatických bunek mléka dojnic indol DNA filtr metodou.

The somatic cells of cow's milk were separated by filtering through nitrocellulose membrane filters (Synpor), pore size 2 to 5 micron. An indole reagent was added to the membrane filters with trapped cells, the mixture was warmed in boiling water bath for 20 min., then cooled, centrifuged (3500 X g, 15 min.), and optical density was measured at 490 nm. The relation 8 micrograms = 1 million cells was used for the conversion of DNA content in sample to the counts of cells. The average variation coefficient of the determination was 4.7% and the coefficient of correlation between the indole DNA filter method and direct microscopy on membrane filters was r = 0.997. Using the indole DNA filter method, the counts of somatic cells can be estimated from milk samples kept frozen or from filters with retained cells kept at 4 degrees C, or (for a short time--3 days) at 25 degrees C. The estimation is not distorted when the milk samples are stabilized with K2Cr2O7 or a mixture of K2Cr2O7 and HgCl2, if they are kept at 4 degrees C for eight days.

[Methods of diagnosing subclinical mastitis in dairy cow mastitis control programs]. / Metody diagnostiky subklinických mastitid v programu tlumení mastitid dojnic.

The number of somatic cells and the isolation of the causative agents of mastitis in quarter, composite, bucket, and bulk tank samples of cow's milk was determined four times during a six-month period. The number of somatic cells in milk samples indicated a degree of mastitis infection and was influenced neither by the year season nor by the length of lactation. At a repeated examination of 28 dairy cows an increased number of somatic cells in milk was found once in 68 udder quarters and with three successive samplings only in 21 quarters. The etiological agents of mastitis were detected once in 31 quarters and three times in succession only in five quarters. The number of cows positive by the number of cells in quarter samples of milk increased from 52.9-58.8% at a single examination to as much as 100% at four examinations. The etiological agents of mastitis were isolated in a single examination in 17.6% of cows and at four examinations in 58.8% of cows. The composite and bucket samples of milk containing 200 to 300 thousand cells per ml are recommended to be considered as mastitis-positive: in 68 to 78% they came from cows having more than 500 thousand cells per ml at least in one quarter sample. The number of cells in a bulk sample was in correlation with the percentage of cows having a positive NK-test (similar to CMT) and positive isolation of S. agalactiae from quarter milk samples.(ABSTRACT TRUNCATED AT 250 WORDS)