Sustained Delivery of Nerve Growth Factor Using a Porous Silicon Film in Rat Pheochromocytoma 12 Cells

Abstract

Source: Rosenberg, M., et al. Designing Porous Silicon Films as Carriers of Nerve Growth Factor. J. Vis. Exp. (2019).

This video demonstrates the use of a degradable nanostructured porous silicon (PSi) film for the sustained release of nerve growth factor (NGF) to support PC12 cell differentiation. NGF is adsorbed onto the film’s pores through electrostatic interactions and is gradually released into the media by diffusion and scaffold degradation. This controlled delivery promotes PC12 cell survival and facilitates neurite growth and branching over an extended period.

Protocol

1. Fabrication of Oxidized porous silicon (PSiO₂) Carriers

1. Cut a Silicon (Si) wafer (single side polished on the <100> face and heavily Boron-doped, p-type, 0.95 mΩ·cm) into 1.5 cm × 1.5 cm samples using a diamond-tipped pen.
2. Oxidize the Si samples in a tube furnace at 400 °C for 2 h in ambient air (heating rate: 25 °C/min, natural cooling).
3. Immerse the Si samples in a solution of aqueous hydrofluoric acid (HF) (48%), double-distilled water (ddH₂O), and ethanol (99.9%) (1:1:3 v/v/v) for 5 min; then rinse the samples with ethanol three times and dry under a nitrogen stream.
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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Acetone	Gadot	830101375
Amphotericin	Biological Industries	03-028-1B
Aqueous HF (48%)	Merck	101513
AZ4533 photoresist	Metal Chem, Inc.	AZ4533
BSA fraction v	MP biomedicals	216006950
BSA solution (10%)	Biological Industries	03-010-1B
Collagen type l	Corning Inc.	354236
Collagenase	Enco	LS004176
Collagen-coated plastic coverslips	NUNC Thermanox	1059846
D-(+)-glucose	Sigma-Aldrich Chemicals	G8170
Dispase-II	Sigma-Aldrich Chemicals	4942078001
Ethanol absolute (99.9%)	Merck	818760
FBS	Biological Industries	04-121-1A
Guanidine-HCl	Sigma-Aldrich Chemicals	G7294
Ham's F-12 nutrient mixture	Thermo Scientific	11765054
HBSS	Thermo Scientific	14185-045
HEPES (1M)	Thermo Scientific	15630-056
HS	Biological Industries	04-124-1A
L-15 medium	Sigma-Aldrich Chemicals	L5520
Laminin	Thermo Scientific	23017015
L-glutamine	Biological Industries	03-020-1A
Murine β-NGF	Peprotech	450-34-20
Normal donkey serum (NDS)	Sigma-Aldrich Chemicals	G9023
Papain	Sigma-Aldrich Chemicals	p-4762
PBS (pH 7.4)			Prepared by dissolving 10 mM Na₂HPO₄, 1.8 mM KH₂PO₄, 137 mM NaCl, and 2.7 mM KCl in double-distilled water (ddH₂O, 18 MΩ·cm).
PBS X10	Biological Industries	02-020-1A
PC12 cell line	ATCC	CRL-1721
Penicillin–streptomycin	Biological Industries	03-032-1B
Poly-L-lysine	Sigma-Aldrich Chemicals	P4832
PrestoBlue reagent	Thermo Scientific	A13261
RPMI medium	Biological Industries	01-100-1A
Si wafer	Siltronix Corp.		Highly-B-doped, p-type, 0.00095 Ω-cm resistivity, <100> oriented
Sodium azide	Sigma-Aldrich Chemicals	S2002
Sodium hydroxide (NaOH)	Sigma-Aldrich Chemicals	S8045
Tannic acid	Sigma-Aldrich Chemicals	403040
Triton X-100	Chem-Impex International Inc.	1279