To determine the absolute viable cell count in the inocula (CFU/ml) and larval homogenates of weighted larvae (CFU/g), 10-fold serial dilutions were prepared in a black 96-well plate (CliniplateTM, Thermo Scientific, Denmark). Fifty microliters of each dilution were plated on Sabouraud agar containing chloramphenicol, incubated at 37°C and counted after 24 and 48 h [27 (link)].
Cliniplate
Manufactured by Thermo Fisher Scientific
Sourced in Denmark
The Cliniplate is a multipurpose laboratory equipment designed for various analytical applications. It serves as a platform for performing assays, reactions, and sample preparation. The Cliniplate provides a reliable and consistent environment for conducting experiments with precision.
Automatically generated - may contain errors
6 protocols using cliniplate
1
Viable Cell Count Determination
2
Fungal Infection Quantification by BLI
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In vitro BLI was performed on the fungal inoculum, and ex vivo BLI on larval homogenates to confirm the relative fungal cell count. Larval homogenates were prepared by placing individual larvae in 600 µl of PBS and homogenizing them using a tissue homogenizer. Ten-fold serial dilutions of the fungal inoculum and larval homogenates were then made in a black 96-well plate (CliniplateTM, Thermo Scientific, Denmark) and 10% d -luciferin potassium salt (1.25 mg/ml in PBS, Promega, USA) was added. The BLI signal was read using an IVIS Spectrum (PerkinElmer, USA) imaging system by acquiring five consecutive images with an exposure time of 30 s (open filter, F/stop1, subject height 0.5 cm, medium binning). Living Image Software (version 4.5.4, PerkinElmer, USA) was used to define regions of interest (ROI) covering each individual well and to calculate the total photon flux (p/s) per well, and peak total fluxes were used for analysis and comparison [27 (link)].
3
Viable Spore Count Determination
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To determine the absolute viable spore counts in the inocula (CFU per milliliter) and larval homogenates (CFU per gram), 10-fold serial dilutions were made in a black 96-well plate (Cliniplate; Thermo Scientific, Denmark). Fifty microliters of each dilution was plated onto Sabouraud agar plates containing chloramphenicol and incubated at 37°C, and spores were counted after 48 h. Experimental data were pooled only if the anticipated inoculum doses were identical.
4
Quantifying Inoculum Spore Counts
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To confirm the relative spore count in the inoculum and larval homogenates, 10-fold serial dilutions were made in a black 96-well plate (Cliniplate; Thermo Scientific, Denmark), and 10% d -luciferin potassium salt (1.25 mg/mL in PBS; Promega, USA) was added. The BLI signal was read using an IVIS Spectrum imaging system (PerkinElmer, USA) by acquiring 5 consecutive images with an exposure time of 30 s (open filter, F/stop1, subject height of 0.5 cm, and medium binning). Living Image software (version 4.5.4; PerkinElmer, USA) was used to define regions of interest (ROIs) covering each well and to calculate the total photon flux (photons per second) per well; peak total fluxes were used for analysis and comparison. Experimental data were pooled only if the confirmed inoculum sizes were identical.
5
Viable Spore Count Determination
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To determine the absolute viable spore counts in the inocula (CFU per milliliter) and larval homogenates (CFU per gram), 10-fold serial dilutions were made in a black 96-well plate (Cliniplate; Thermo Scientific, Denmark). Fifty microliters of each dilution was plated onto Sabouraud agar plates containing chloramphenicol and incubated at 37°C, and spores were counted after 48 h. Experimental data were pooled only if the anticipated inoculum doses were identical.
6
Quantifying Inoculum Spore Counts
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To confirm the relative spore count in the inoculum and larval homogenates, 10-fold serial dilutions were made in a black 96-well plate (Cliniplate; Thermo Scientific, Denmark), and 10% d -luciferin potassium salt (1.25 mg/mL in PBS; Promega, USA) was added. The BLI signal was read using an IVIS Spectrum imaging system (PerkinElmer, USA) by acquiring 5 consecutive images with an exposure time of 30 s (open filter, F/stop1, subject height of 0.5 cm, and medium binning). Living Image software (version 4.5.4; PerkinElmer, USA) was used to define regions of interest (ROIs) covering each well and to calculate the total photon flux (photons per second) per well; peak total fluxes were used for analysis and comparison. Experimental data were pooled only if the confirmed inoculum sizes were identical.
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