Table 4.

Selected chemical reporters for mycobacterial glycans

Target	Metabolite	Compound	Reporter group (X)	Bioorthogonal reaction(s)a	Proposed route	Speciesb	Applicationsc	Synthesis	Notesb	Refd
TMM and/or TDM	Trehalose	FITC-Tre 93 (Fig. 35)	Fluorescein	--	Extracellular	Broad (M)	In vitro and in vivo imaging (infected macrophages)	Chemical; comm. available	Contains unnatural anomeric methyl group	264
	Trehalose	FlTre series 94–97 (Fig. 35)	Fluorescein	--	Extracellular	Broad (M)	In vitro imaging; membrane fluidity and inhibitor characterization	Chemical	Lack of unnatural anomeric methyl group enhances incorporation	265
	Trehalose	DMN-Tre 98 (Fig. 35)	DMN	--	Extracellular	Broad (M)	In vitro and in vivo imaging (infected macrophages); selective detection of M. tuberculosis; inhibitor characterization	Chemical	Fluorogenic	266
	Trehalose	TreAz series 99–102 (Fig. 35)	Azide	CuAAC; SPAAC	Intracellular	Broad (M)	In vitro and in vivo imaging (infected host organism); inhibitor characterization	Chemo-enzymatic; chemical; 6-TreAz is comm. available	2-, 4-, 6-TreAz label via LpqY-SugABC; 3-TreAz labels via Ag85; 6-TreAz only labels TMM	267
	Trehalose	6-LactoTreAz 104 (Fig. 35)	Azide	SPAAC	Intracellular	M. smegmatis (M)	In vitro imaging	Chemo-enzymatic	Inversion of 4-position enhances stability toward trehalase	268
	Trehalose	18F-2-FDTre 105 (Fig. 35)	18F	--	Intracellular	M. smegmatis (M)	In vitro tracing of trehalose uptake	Chemo-enzymatic	Potential in vivo PET imaging probe	269,270
	TMM	N-AlkTMM-C7 121 (Fig. 38)	Alkyne	CuAAC	Extracellular	Broad (M)	In vitro imaging	Chemical	Only labels TDM	271
	TMM	N-x-AlkTMM-C15 122 (Fig. 38)	Alkyne and diazirine	CuAAC	Extracellular	Broad (M)	Identification of mycolate–protein interactions	Chemical	Diazirine allows protein photo-cross-linking	272
AGM and TDM	TMM	O-AlkTMM-C7 113 (Fig. 38)	Alkyne	CuAAC	Extracellular	Broad (M)	In vitro imaging; protein profiling	Chemical	Also labels O-mycoloylated proteins in C. glutamicum	271
	TMM	O-AlkTMM-C23 115 (Fig. 38)	Alkyne	CuAAC	Extracellular	C. glutamicum (related to M)	In vitro imaging	Chemical	Possesses more-native mycolate chain	273
	TMM	O-AzTMM-C10 116 (Fig. 38)	Azide	SPAAC; CuAAC	Extracellular	M. smegmatis (M), C. glutamicum (related to M)	In vitro imaging	Chemical	Also labels O-mycoloylated proteins in C. glutamicum	274
	TMM	O-TCO-TMM-C10 117 (Fig. 38)	Trans-cycooctene	Tetrazine ligation	Extracellular	M. smegmatis (M), C. glutamicum (related to M)	In vitro imaging; rapid detection of mycobacteria	Chemical	Also labels O-mycoloylated proteins in C. glutamicum	274
	TMM	O-FITC-TMM-C10 119 (Fig. 38)	Fluorescein	--	Extracellulare	M. smegmatis (M), C. glutamicum (related to M)	In vitro imaging	Chemical	Labeling of O-mycoloylated protein was inconsistent with other TMM reporters	274
	TMM	O-DBF-TMM 120 (Fig. 38)	Dibromo-fluorescein	--	Extracellular	C. glutamicum (related to M)	In vitro imaging using transmission electron microscopy	Chemical	DBF is a singlet oxygen-generating fluorophore	275
Ag85 enzyme	TMM	QTF 125 (Fig. 40)	FRET pair	--	Extracellular	M. smegmatis (M), C. glutamicum (related to M)	Detection and imaging of Ag85 hydrolytic activity	Chemical	Fluorogenic probe of enzyme activity	276
Tdmh enzyme	TDM	FRET-TDM 128 (Fig. 42)	FRET pair	--	Extraceullar	M. smegmatis (M), M. tuberculosis (M)	Detection of Tdmh hydrolytic activity	Chemical	Fluorogenic probe of enzyme activity	277
AG, other arabinans	Arabinose	5-AraAz 133 (Fig. 45)	Azide	SPAAC	Intracellulare	M. tuberculosis (M)	In vitro imaging and in vivo imaging (macrophages infected with pre-labeled bacteria)	Chemical	Labels M. tuberculosis cells but route and target are unknown	278
	DPA	2-AzFPA 135 (Fig. 46)	Azide	SPAAC	Extracellular	M. smegmatis (M), C. glutamicum (related to M)		Chemical	Lipid-linked Araf derivative	279
	DPA	5-AzFPA 137 (Fig. 46)	Azide	SPAAC	Extracellular	M. smegmatis (M), C. glutamicum (related to M)	In vitro and in vivo imaging (macrophages infected with pre-labeled bacteria)	Chemical	Lipid-linked Araf derivative	279

^aNot applicable for fluorescent reporters operating via one-step incorporation (--).

^bGram classification: GN, Gram-negative; GP, Gram-positive; M, mycobacteria.

^cIn vitro refers to bacterial labeling experiments performed in broth culture; in vivo refers to bacterial labeling experiments performed in a host cell or organism.

^dInitial publication(s) referenced. See text for references for follow-up studies and applications.

^eLimited evidence available to support proposed route of incorporation and/or identity of labeled target (see text).