Decalcified stones were dehydrated through a series of graded ethanol
concentrations and then embedded in paraffin. Paraffinized and unstained
(label-free) sections of stones were imaged by fluorescence on a Leica TCS SP8
imaging system; and multiphoton excitation at 910 nm for second harmonic
generation (SHG) imaging was used for collagen detection (Ferkowicz et al., 2021 (link)). Subsequent staining of human
renal tissue and demineralized stone sections were deparaffinized, permeabilized
with 0.05% Triton X-100 and stained using 7-aminoactinomycin D (7-AAD) (Thermo
Fisher Scientific, Itasca, Illinois, USA) or
4’,6-diamindino-2-phenylindole (DAPI) (Vector Laboratories, Burlingame,
CA, USA) for DNA detection. Additional sections were stained by routine
hematoxylin & eosin (H&E) methods. Observations were all done in
triplicate, except where noted.
concentrations and then embedded in paraffin. Paraffinized and unstained
(label-free) sections of stones were imaged by fluorescence on a Leica TCS SP8
imaging system; and multiphoton excitation at 910 nm for second harmonic
generation (SHG) imaging was used for collagen detection (Ferkowicz et al., 2021 (link)). Subsequent staining of human
renal tissue and demineralized stone sections were deparaffinized, permeabilized
with 0.05% Triton X-100 and stained using 7-aminoactinomycin D (7-AAD) (Thermo
Fisher Scientific, Itasca, Illinois, USA) or
4’,6-diamindino-2-phenylindole (DAPI) (Vector Laboratories, Burlingame,
CA, USA) for DNA detection. Additional sections were stained by routine
hematoxylin & eosin (H&E) methods. Observations were all done in
triplicate, except where noted.