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

Table 2

Magnetic separation methods used for the detection of Mycobacterium avium subsp. paratuberculosis cells in feces.

Method	Type of beads	Coating ligand	Type of matrix	Artificially contaminated	Preparation of sample	Initial volume of sample	Initial number of MAP [MAP/mL]	Limit of Detection	Sensitivity [%]	Reference
IMS	BioMag goat anti-rabbit IgG	Rabbit polyclonal anti-MAP antibodies	Bovine	Yes	MAP aliquots added to 200 mg of feces, resuspended in PBS (2 mL), mixed on a rotating platform, and centrifuged, clear upper phase transferred to a tube with beads	200 mg	10^6 to 10^0	10 or fewer MAP/200 mg	100	[35]
PMS	MyOne Tosylactivated Dynabeads	Biotinylated peptides (aMp3, aMptD)	Bovine	No	Sample mixed with 4 mL of sterile water and centrifuged low-speed, 1 mL of supernatant used for PMS	1 g	n. a.	from 6 to 41,1 PFU/g	n. a.	[19]
			Bovine	No	No preparation	2 g	n. a.	n. a.	n. a.	[37]
			Bovine	Yes	Dilution of autoclaved bovine feces in sterile water 1 : 5; serial 10-fold dilutions of MAP prepared in MB containing 2 mmol/L CaCl2 and added (100 µL) to 900 µL aliquots of fecal suspension	1 mL		from 3 × 10^2 to 6 × 10^8 phage mL^−1	n. a.	[17]

BioMag goat anti-rabbit immunoglobulin G [IgG] with rabbit polyclonal anti-MAP antibodies (Polysciences, Inc., Warrington, Pa.); the number of PFU mL⁻¹ used to spike the samples was determined by subjecting the 10⁻³, 10⁻⁴, and 10⁻⁵ spiking dilutions to the optimized phage assay and counting the plaques produced. In the final assay format, a consistent number of D29 seed phages (10⁸ PFU mL⁻¹) was added to the samples; IMS = immunomagnetic separation; MAP = Mycobacterium avium subsp. paratuberculosis; MB = Middlebrook 7H9 broth; n. a. = not available; PBS = phosphate-buffered saline; PFU = plaque-forming units; PMS = peptide-mediated separation.