Magnetic Separation Methods for the Detection of Mycobacterium avium subsp. paratuberculosis in Various Types of Matrices: A Review

Table 5

Magnetic separation methods used for the detection of Mycobacterium avium subsp. paratuberculosis DNA in milk.

Type of kit	Type of milk	Artificially contaminated	Preparation of sample	Initial volume of sample [mL]	Initial number of MAP [MAP/mL]	Number of copies detected	Analytical sensitivity [MAP/mL]	Sensitivity of detection [%]	Reference
Adiapure	Bovine raw	Yes	No preparation	10	10^6–10^7	30 copies of IS900/mL	n. a.	100, 90, 85, 25	[41]
EZ1 DNA Tissue Kit	Bovine BTM	No	Milk centrifuged, cream retained, pellet incubated (37°C), centrifuged; a mixture of 1000 µL lysis buffer, 2 µL proteinase K used to dissolve the cream fraction, dissolved cream pooled with the pellet, mixture transferred to bead beating tubes (600 µL of beads), tubes incubated (56°C), shaken, centrifuged, 400 µL of the liquid phase processed with kit and BioRobot	10	10^0–10^4	n. a.	100	n. a.	[42]

The combined Adiapure-Adiavet MAP DNA extraction and detection kit with magnetic beads; artificially contaminated with naturally infected feces (10⁶–10⁷ MAP); 10 g thawed feces diluted in 90 mL sterile distilled water; sensitivity of detection was 100, 90, 85, and 25% for respective MAP concentrations of 300, 30, 3, and 0.3 copies of IS900/mL⁻¹; EZ1 DNA Tissue Kit with magnetic beads (Qiagen, Hilden, Germany) automatically processed in the BioRobot® EZ1 workstation (Qiagen); for assessment of analytical sensitivity, quantification of MAP and the testing of various parameters during the development of the extraction method artificially contaminated milk was used; auto = automatically processed; BTM = bulk tank milk; MAP = Mycobacterium avium subsp. paratuberculosis; n. a. = not available.