TABLE 1.

List of Primary and Secondary Antibodies Used for Fluorescence Staining of Biotinylated Dextran Amine (BDA), Vesicular Transporter Proteins (VTPs) and Postsynaptic Proteins (PSPs)

	1ry AB (overnight)							2ry AB (3 hours)		SA (3 hours)
Series	Antigen	Host	Dilution	Source	Cat. #	Immunogen	Specificity	IgG (Dky)17,18	Dilution	SA17	Dilution	n19
Double BDA VTP	VAChT	Gt	1:5000	Chemicon1	AB1578	Synthetic peptide corresponding to C-terminus of cloned rat VAChT6 (CSPP GPFDGCEDDYNYYSRS)7	By WB the AB recognizes a ≈65–70 kD band corresponding to VAChT protein4	Anti-Gt-Cy3	1:800	SA-Cy2	1:800	8
	VAChT	Rb	1:1000	Sigma2	V5387	Synthetic peptide corresponding to AA 512–530 of C-terminus of cloned rat VAChT (K- SPPGPFDGCEDDYNYYSRS)8	By WB the AB recognizes a ≈67–70 kD band, corresponding to VAChT protein7	Anti-Rb-Cy3	1:800	SA-Cy2	1:800	9
	VGluT1	Rb	1:1000	Gift RHE3	–	Synthetic peptide corresponding to 68 last AA of C-terminus of rat BNPi (VGluT1)9	By WB the AB recognizes a ≈62 kD band from rat brain9	Anti-Rb-Cy3	1:800	SA-Cy2	1:800	4
	VGluT2	Rb	1:5000	Gift RHE	–	Synthetic peptide corresponding to 64 last AA of C-terminus of rat DNPi (VGluT2)10	By WB the AB recognizes a ≈50–62 kD band from rat brain10	Anti-Rb-Cy3	1:800	SA-Cy2	1:800	14
	VGluT3	GP	1:1000	Chemicon	AB5421	Synthetic peptide from cloned rat VGluT3 protein11 (AFEGEEPLSYQNEEDFSETS)7	The AB labels VGluT3+ cells and fibers, in agreement with other VGluT3 antisera11	Anti-GP-Cy3	1:800	SA-Cy2	1:800	7
	VGAT	Rb	1:250	Chemicon	AB5062P	Synthetic peptide corresponding to a 17 AA peptide near C- terminus region of rat VGAT12 (VHSLEGLIEAYRTNAED)7	By WB the AB recognizes a band at ≈55–60 kD12	Anti-Rb-Cy3	1:800	SA-Cy2	1:800	14
Double VTP VTP	VAChT/VGluT2	Gt Rb	1:5000 1:5000	Chemicon Gift RHE	AB1578 -	(above) (above)	(above) (above)	Anti-Gt-Cy3 Anti-Rb-Cy2	1:800 1:200			4
	VAChT/VGAT	Gt Rb	1:5000 1:250	Chemicon Chemicon	AB1578 AB5062P	(above) (above)	(above) (above)	Anti-Gt-Cy3 Anti-Rb-Cy2	1:800 1:200			4
	VGAT/VGluT2	Rb GP	1:250 1:5000	Chemicon Chemicon	AB5062P AB5907	(above) Synthetic peptide from cloned rat VGluT2 protein13 (VQESAQDAYSYKDRDDYS)7	(above) The AB gives labeling in agreement with other antisera to VGluT213	Anti-Rb-Cy2 Anti-GP-Cy3	1:200 1:800			4
Triple BDA VTP PSP	VGluT2/PSD-95	Rb Ms	1:5000 1:100	Gift RHE ABR4	- MA1-045	(above) Purified recombinant of rat PSD- 9514	(above) The AB detects post synaptic density 95kD in rat brain. By WB it recognizes a ≈95 kDa band14	Anti-Rb-Cy5 Anti-Ms-Cy3	1:800 1:800	SA-Cy2 SA-Cy2	1:800 1:800	3
	VGAT/Geph	Rb Ms	1:250 1:100	Chemicon SY-SY5	AB5062P 147 011	(above) Purified rat gephyrin15	(above) By WB the AB recognizes a ≈93 kD band. It detects a N-terminus epitope16	Anti-Rb-Cy5 Anti-Ms-Cy3	1:800 1:800	SA-Cy2 SA-Cy2	1:800 1:800	3

¹Chemicon: Chemicon International, Temecula, CA

²Sigma: Sigma, St. Louis, MO

³Gift from Edwards, R.H. and Fremeau, R.T. Jr.

⁴ABR: Affinity BioReagents, Golden, CO.

⁵SY-SY: Synaptic Systems, Göttingen, Germany.

⁶http://www.chemicon.com/Product/ProductDataSheet.asp?ProductItem=AB1578

⁷Supplied by Chemicon on request

⁸http://www.sigmaaldrich.com/sigma/datasheet/v5387dat.pdf

⁹Bellocchio et al., 1998

¹⁰Fremeau et al., 2001

¹¹http://www.chemicon.com/Product/ProductDataSheet.asp?ProductItem=AB5421

¹²http://www.chemicon.com/Product/ProductDataSheet.asp?ProductItem=AB5062P

¹³http://www.chemicon.com/product/productdataSheet.asp?ProductItem=AB5907

¹⁴http://www.bioreagents.com/index.cfm/fuseaction/products.print/Product/MA1-045

¹⁵http://www.sysy.com/gephyrin/gephy_fs.html

¹⁶Pfeiffer et al., 1984

¹⁷Jackson Immuno Research Laboratories, West Grove, PA.

¹⁸For multiple labeling (ML) with minimal cross-reactivity (min X) to other species.

¹⁹n, number of cases (each case referring to an injection site and series from the same side of the brain, thus 1 or 2 per brain).

AA, amino acid; AB, antibody; BNPi, brain specific Na⁺-dependent phosphate transporter; Cat., catalog; Cy2, cyanine; Cy3, indocarbocyanine; Cy5, indodicarbocyanine; Dky, donkey; DNPi, differentiation-associated Na⁺-dependent phosphate transporter; Geph, gephyrin; GP, guinea pig; Gt, goat; Ms, mouse (monoclonal); Rb, rabbit; SA, streptavidin; WB, western blot.