Target Peptide Detection and Measurement Using a Capacitive Immunoprobe Biosensor

Abstract

Source: Kluge, N. et al., Time-Resolved In Vivo Measurement of Neuropeptide Dynamics by Capacitive Immunoprobe in Porcine Heart. J. Vis. Exp.(2022).

This video demonstrates a method for detecting and quantifying peptide transmitters using a capacitive immunoprobe biosensor. The functionalized electrode, coated with antibodies, interacts with peptides of interest, leading to a signal change that allows precise measurement and detection of peptides.

Protocol

1. Capacitive immunoprobe fabrication and functionalization

1. Cut a 25 cm length of perfluoroalkoxy (PFA)-coated platinum wire (see Table of Materials) and strip approximately 5 mm of PFA coating from one end using a scalpel, being careful not to cut into the platinum wire.
2. Insert the stripped end of the platinum wire into a gold-plated 1 mm male connector pin and crimp the connector pin teeth around the stripped end of the platinum wire using needle-nose pliers (see Table of Materials).
3. Solder the platinum wire to the gold-plated connector pin. Be careful not to use an excessiv....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
AgCl disc electrode	Warner Instruments (Holliston, MA)	64-1307
Anti-NPY monoclonal antibody	Abcam, (Cambridge, MA)	ab112473
Custom multichannel amplifier/ 1 MΩ feedback resistor multichannel headstage	NPI Electronic, (Tamm, Germany)	NA	Based on NPI VA-10M multichannel amplifier
Dopamine HCl	Sigma Aldrich (St. Louis, MO)	H8502-10G
Gold-plated male connector pin	AMP-TE Connectivity (Amplimite)	6-66506-1
HEKA LIH 8+8 analog-to-digital/digital-to-analog device	HEKA Elektronik, (Holliston, MA)	NA
Igor Pro data acquisition software, v. 7.08	WaveMetrics, (Lake Oswego, OR)		Software driving command potential and data acquisition was custom written
Masterflex L/S Standard Digital peristaltic pump	Cole Palmer, (Vernon Hills, IL)
PFA-coated platinum wire	A-M Systems, (Sequim, WA)	773000	0.005" bare diameter, 0.008" coated diameter
Silicone elastomer	World Precision Instruments (Sarasota, FL)	SYLG184
Synthetic porcine NPY peptide	Bachem (Torrance, CA)	4011654