A follow-up FP assay was performed in a half-area black plate (Corning, Corning, New York, 3686). Fifty microliters of 125 nM MSUT2 and 10 nM FAM-RNA (ordered from IDT) in PBS (137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, and 1.8 mM KH2PO4) was transferred using an Integra (Hudson, NH) Viaflo with a 96/50 μL head. Two microliters of compound was transferred for a final concentration of 10 μM. Plates were incubated for 20 min at room temperature away from light and read using a Cytation 5 (BioTek, Winooski, VT) with a preconfigured green polarization filter cube (Biotek, 8040561) at excitation 485/20 and emission 528/20, a dichroic mirror at 510 nm, and a read height of 10 mm. FP was calculated by first subtracting the background from a buffer-only control well and then using the equation to determine polarization (P).
Half area black plate
Manufactured by Corning
The Half-area black plate is a laboratory equipment item designed to provide a consistent dark surface for various scientific experiments and applications. It serves as a neutral background for visual and photographic observations, helping to enhance contrast and visibility. The product's core function is to offer a standardized, predictable dark surface for use in laboratory settings.
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2 protocols using half area black plate
1
Fluorescence Polarization Assay for MSUT2
2
Fluorescent Protein Synthesis Quantification
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The fluorescence
intensity of the synthesized fluorescent proteins was measured by
the multiwell plate fluorometer (Synergy HTX, BioTek, Winooski, VT).
Five microliters of the cell-free synthesized fluorescent protein
and 45 μL of Milli-Q water were mixed in a 96-well half-area
black plate (Corning Incorporated, Corning, NY). The plate was mixed
in the plated reader orbitally at a medium speed for 15 s and read
at a height of 1.5 mm with a gain of 48. The excitation and emission
spectra are 485 and 528 nm, respectively. The cell-free synthesized
protein was visualized by Coomassie blue staining after protein gel
electrophoresis using precasted 4–12% bis–tris gradient
gel (Invitrogen, Waltham, MA) (Figure S6 ).
intensity of the synthesized fluorescent proteins was measured by
the multiwell plate fluorometer (Synergy HTX, BioTek, Winooski, VT).
Five microliters of the cell-free synthesized fluorescent protein
and 45 μL of Milli-Q water were mixed in a 96-well half-area
black plate (Corning Incorporated, Corning, NY). The plate was mixed
in the plated reader orbitally at a medium speed for 15 s and read
at a height of 1.5 mm with a gain of 48. The excitation and emission
spectra are 485 and 528 nm, respectively. The cell-free synthesized
protein was visualized by Coomassie blue staining after protein gel
electrophoresis using precasted 4–12% bis–tris gradient
gel (Invitrogen, Waltham, MA) (
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