Ammonium persulfate

Round microscopy cover slides were washed with 70% ethanol and 0.1 m NaOH, covered with 3‐aminopropyltrimethoxysilane (3‐APTS, Sigma) for 3 min for activation, incubated for 30 min in 0.5% glutaraldehyde (Sigma), and washed in sterile MilliQ water. Polyacrylamide solutions containing acrylamide monomers (Sigma), crosslinker N,N‐methylene‐bis‐acrylamide (Sigma) and PBS in different concentrations were prepared to create different Young's modulus (0.5, 2, 4.5, 10, 20 and 115 kPa) as previously described [27 (link)]. 5 μL of 10% ammonium persulfate (Sigma) and 0.75 μL N,N,N′,N′‐tetramethylethylenediamine (Sigma) were added into 0.5 mL mixtures, and one drop of the mixture was placed on rain repellent‐treated microscopy slides, and the activated cover slides were placed on top. After polymerisation (3–10 min), the cover slides with polyacrylamide gels were washed with PBS. Next, the cover slides were treated with 1 mg·mL⁻¹N‐sulfosuccinimidyl‐6‐(4′‐azido‐2′‐nitrophenylamino) hexanoate (Sigma) and exposed to ultraviolet (UV) light to allow subsequent collagen‐I binding. The cover slides were incubated at room temperature with 10 μg·mL⁻¹ rat‐tail collagen‐I (Sigma) for 3 h, washed with PBS and placed in UV light for sterilisation. Wells containing cover slides were seeded with SKUT1 cells or primary SMC, uLMS, LM or MM cells.

To prepare polyacrylamide gels, glass coverslips were washed twice with 70% ethanol, activated with 0.1 M NaOH, and silanized with (3-aminopropyl) trimethoxysilane (Sigma). Then the glass coverslips were treated with 0.5% glutaraldehyde (Sigma) for 30 min and washed extensively with MilliQ water. Mixtures of MilliQ water, acrylamide monomers (Sigma), and crosslinker N,N-methylene-bis-acrylamide (Sigma) were prepared according to previously determined formulations^{62 ,63 (link)}. For the polymerization reaction, 5 μl of 10% ammonium persulfate (Sigma) and 0.75 μl N,N,N′,N′-tetramethylethylenediamine (Sigma) were added into 0.5 ml mixtures. To functionalize gels with collagen, gels were first treated with 1 mg/ml N-sulfosuccinimidyl-6-(4′-azido-2′-nitrophenylamino) hexanoate (Sigma), which was activated by ultraviolet (UV) light. Finally, gels were incubated with 10 μg/ml collagen I from rat tail (Sigma) or human collagen VI (Rockland) for 3 h at room temperature (RT) and UV-sterilized prior to cell seeding.

N-isopropylacrylamide (NIPA), N-(3-aminopropyl)methacrylamide hydrochloride (APMA), N,N′-methylenebisacrylamide (BIS), ammonium persulfate (APS), N,N,N′,N′-tetramethylethylenediamine (TEMED), gallic acid (GA), N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide (EDC), N-hydroxysuccinimide (NHS), dimethylformamide (DMF), triethylamine (TEA), sodium nitrate (NaNO₃), chloric acid (HCl), sodium hydroxide (NaOH), and D₂O were purchased from Aldrich. All chemicals were used as received, except for NIPA, which was recrystallized from the toluene-hexane mixture (90:10 v/v). All solutions were prepared using high purity water obtained from a Hydrolab purification system (water conductivity: 0.05 μS·cm⁻¹).

N-(3-Aminopropyl)methacrylamide
hydrochloride (APMA), N-isopropylacrylamide (NIPA), N,N′-methylenebis(acrylamide) (BIS), N,N,N',N'-tetramethylethylenediamine (TEMED),
ammonium persulfate
(APS), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)
diammonium salt (ABTS), and ascorbic acid were purchased from Aldrich.
Sodium nitrate (NaNO₃) and sodium hydroxide were purchased
from POCh. NIPA was purified using recrystallization from a mixture
of toluene/hexane (30:70, v/v). All other chemicals were used as received.
The solutions were prepared using high-purity water obtained from
a Milli-Q Plus/Millipore purification system (water conductivity of
0.056 μS cm^–1).

NVF (98%), potassium tert-butoxide (PTB, 95%), bis(2-bromoethyl) ether (BBE, 95%), dicyclohexyl-18-crown-6
(98%), anhydrous tetrahydrofuran (THF, 99.9%), chloroform (CHCl₃, 98%), ethanol (99.9%), azoisobutyronitrile (AIBN, 98%),
and poly(1-vinyl-pyrrolidone-co-vinyl acetate) (poly(VP-co-VA)) (average M_w ∼
50,000) were all purchased from Aldrich and used as received. EA (99%),
MAA (99%), glycidyl methacrylate (GMA, 97%), divinylbenzene (DVB,
80%), NaOH (97%), ammonium persulfate (APS, 98%), sodium dodecyl sulfate
(SDS), N,N,N′,N′-tetramethyl ethylenediamine (TEMED, 99%), dipotassium
phosphate (K₂HPO₄, 97%), and methylene violet
(3RAX) were also purchased from Aldrich and used as received. All
water was of ultra-high-purity deionized quality.

β-Cyclodextrin (β-CD),
aminoferrocene, N-isopropylacrylamide (NIPA), N,N′-methylenebis(acrylamide) (BIS),
rhodamine B, and ammonium persulfate (APS) were purchased from Aldrich.
Sodium nitrate (NaNO₃), hydrochloric acid (HCl), and potassium
chloride (KCl) were purchased from POCh. All reagents were used as
received, except NIPA, which was purified with recrystallization from
a toluene/hexane mixture (3:7, v/v). All solutions were prepared using
high-purity water from a Milli-Q Plus/Millipore purification system
(conductivity 0.055 μS·cm^–1). 6-Amino-β-CD
was synthesized according to the procedure described earlier.^{52 (link)}

First, 9.80 mg of MSA (Sigma-Aldrich), 9.50 mg of EDSA (Tokyo Chemical Industry), 11.8 mg of BDSA (Ark Pharm) and 15.7 mg of BPDSA (Tokyo Chemical Industry) were each individually dissolved in 5 ml of deionized water in 20 ml capped vials. Then, 7.00 μl of Py (Tokyo Chemical Industry) was added to each vial. The mixtures of Py and acid were stirred for 2 h at ∼0 °C to prepare the TMCPs. The substrates (glass slides, indium tin oxide-coated glass slides, poly(ethylene terephthalate) films or graphite foils) were successively ultrasonicated with deionized water, acetone (Sigma-Aldrich) and isopropyl alcohol (Aldrich) for 15 min each. Clear substrates were vertically immersed in each vial. The washed substrates were then dried in an oven at 60 °C, and their surfaces were treated by means of combined exposure to ultraviolet and ozone. The in situ chemical oxidative polymerization of P(Py:MSA) and P(Py:DSA:Py) was induced on the substrates at ∼0 °C by adding 5 ml of a 0.10 mmol ammonium persulfate (Aldrich) aqueous solution. After 12 h, uniform P(Py:MSA) and P(Py:DSA:Py) films were obtained on the substrates. The films were rinsed with deionized water and ethanol (Aldrich) to remove any remaining salts and unreacted monomers. The washed films were then dried in a vacuum oven at 80 °C overnight.