Cpd 030 critical point dryer

Chemicals
were bought from Sigma-Aldrich, unless differently specified. Samples
(n = 4) were rinsed with PBS and fixed with 4% paraformaldehyde
for 30 min at RT. After washing with PBS, samples (n = 2) were dehydrated with a sequential series of 70–80–90–100%
ethanol solutions, 30 min per step. After dehydration, samples were
dried with a critical point dry setup (CPD 030 Critical Point Dryer,
Leica) and then gold-sputtered at 40 mA and 100 mTorr for 30 s. Cell
morphology was investigated using a Philips XL30 ESEM-FEG SEM at 10
kV and a working distance of 10 mm.
For other polymeric discs
(n = 2), attached cells were permeabilized and blocked
with TBP buffer (0.1% Triton X-100, 0.5% bovine serum albumin in PBS)
overnight at 4 °C. Cells were stained with monoclonal antiactin,
α-smooth muscle (1:200) conjugated with goat antimouse Alexa
Fluor 488 (Invitrogen, 1:1000) as the secondary antibody, Phalloidin-Texas
red (Molecular Probes, 1:100), and 4′,6-diamidino-2-phenylindole
(DAPI) (1:100) with three times washing steps in between. A Nanozoomer
slide scanner equipped with a 40× objective (Hammamatsu) was
used to acquire images.

Briefly, samples for SEM were fixed in 2% glutaraldehyde in cacodylate buffer, followed by rinsing in buffer and dehydration in a graded ethanol series. The samples were critical point dried in a Bal-tec CPD030 critical point dryer, mounted on aluminum stubs, gold coated in a Leica ACE200 sputter coater, and examined in an FEI XL30 SEM.

As the first step of self-rolling, a layer of photoresist (AZ-5214, Micro-chemicals GmbH, Germany) with about 1 μm thickness was spin-coated (4000 RPM, 30 s, KW-4A, SIYOUYEN, China) and etching windows were defined by photolithography (HEIDELBERG, UPG501). After development, the window was exposed and etched by reactive ion etching (RIE, Trion T2) for 60 s (30 sccm CF₄ and 30 sccm Ar flow, 300 mTorr chamber pressure, and 100 W etching power). The photoresist layer was removed by ultrasonication in ethanol (99.7%) lasting for 30 s. For the rolling process, patterned VO₂ NMs were released from the substrate by using 40% HF (hydrofluoric acid) solution at room temperature for 5 min. Due to the high selectivity, the property of the VO₂ NMs can hardly be influenced by this etching process. Finally, critical point drying (Leica CPD030 Critical Point Dryer) was applied to dry the rolled-up structures, avoiding structural collapse.