Improved bioluminescent assay of somatic cell counts in raw milk.
Somatic cell count (SCC) in milk is considered to be a valuable indicator of cow mastitis. For assessment of SCC in milk, the bioluminescent assay based on determination of ATP from somatic cells ([ATPsom]) in milk was proposed earlier. However, this assay is still not widely used in practice owing to lower reliability compared with conventional methods such as direct microscopy and flow cytometry. We revised the bioluminescent SCC assay and developed a simple protocol based on determination of the total non-bacterial ATP concentration in milk. It was shown that the novel ATP-releasing agent Neonol-10 (oxy-ethylated iso-nonyl phenol) has superior performance providing 100% lysis of somatic cells while not disrupting bacterial cells of milk at a concentration of 1.5% w/w. There was high correlation (R2=0.99) between measured bioluminescence and SCC as measured by direct microscopy. The observed detection limit of the bioluminescent milk SCC assay was as low as 900 cell/ml, time of analysis was 2-3 min per sample. The proposed method has high potential for on-site mastitis diagnostics.
Frundzhyan
Valery G
VG
Chemistry Department, Lomonosov Moscow State University, Moscow 119991, Russia.
Parkhomenko
Inna M
IM
Brovko
Lubov Y
LY
Ugarova
Natalia N
NN
D000255:Adenosine Triphosphate / Q000032:analysis; D000818:Animals; D002417:Cattle; D002452:Cell Count / Q000379:methods*; D002477:Cells / Q000187:drug effects / Q000201:enzymology; D003902:Detergents / Q000494:pharmacology; D008163:Luminescent Measurements / Q000379:methods*; D008414:Mastitis, Bovine / Q000175:diagnosis; D008892:Milk / Q000737:chemistry / Q000166:cytology* / Q000201:enzymology
D003902:Detergents; D000255:Adenosine Triphosphate