Mouse infections were confirmed by luciferase measurement of mouse fecal samples following a previously reported protocol (43 (link)). Briefly, 20 mg of fecal sample was weighed into a 1.5-ml microcentrifuge tube and homogenized in 1 ml of lysis buffer (50 mM Tris-HCl, 10% glycerol, 1% Triton-X, 2 mM dithiothreitol [DTT], 2 mM EDTA) using 10 to 15 glass beads (3 mm) and a vortex mixer for 1 min, followed by clarification of lysate by brief centrifugation. One hundred microliters of lysate was mixed with an equal volume of NanoGlo luciferase buffer, prepared with a 1:50 dilution of the substrate (Promega). Three technical replicates per sample were conducted. Luminescence was measured in a Synergy HT Luminator (BioTek).
Nano glo luciferase buffer
Manufactured by Promega
The Nano-Glo luciferase buffer is a component used in luminescence-based assays. It provides the necessary reagents to detect and quantify luciferase activity in samples. The buffer contains the substrate required for the luciferase reaction, enabling the measurement of luminescent signal.
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Lab products found in correlation
4 protocols using nano glo luciferase buffer
1
Fecal Luciferase Measurement for Mouse Infection
2
Luminescent Protein Quantification
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The supernatants were cultured in a mixture of Nano-Glo luciferase buffer and the substrate (catalog no. N1120; Promega) (100:1) at room temperature for 15 min. All of the liquid was transferred to an opaque plate, and the chemiluminescence value was determined using a GloMax 96 luminometer (Promega).
3
Luminescent Protein Quantification
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The supernatants were cultured in a mixture of Nano-Glo luciferase buffer and the substrate (catalog no. N1120; Promega) (100:1) at room temperature for 15 min. All of the liquid was transferred to an opaque plate, and the chemiluminescence value was determined using a GloMax 96 luminometer (Promega).
4
Megalin-mediated NL-D3 Uptake Assay
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Parental HEK293 EBNA cells or HEK293 EBNA cells stably expressing megalin minireceptor (MmR4) were kindly provided by Prof. Thomas Willnow (Max Delbruck Centre for Molecular Medicine, Berlin). The cells were maintained at 37˚C in 5% CO2 in DMEM containing 10% fetal bovine serum, penicillin (100 U/ml), streptomycin (0.1 mg/ml), and 0.35 mg/ml G418 without (parental) or with (MmR4 cells) an additional 1 µg/ml puromycin.
For NL-D3 uptake, cells were washed with PBS, then serum-free DMEM before preincubating in serum-free DMEM for 30 min at 37˚C. Cells were then incubated in serumfree DMEM containing NL-D3 at 1 µg/ml for 30 min at 37˚C, washed 3x in PBS, and then lysed in Nano-Glo luciferase buffer (Promega) for assessment of luciferase activity.
For NL-D3 uptake, cells were washed with PBS, then serum-free DMEM before preincubating in serum-free DMEM for 30 min at 37˚C. Cells were then incubated in serumfree DMEM containing NL-D3 at 1 µg/ml for 30 min at 37˚C, washed 3x in PBS, and then lysed in Nano-Glo luciferase buffer (Promega) for assessment of luciferase activity.
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