Chromate 4300 microplate reader

Manufactured by Awareness Technology

Sourced in United States

The Chromate 4300 Microplate Reader is a laboratory instrument designed for high-throughput absorbance measurements. It is capable of reading microplates with multiple wells, providing efficient data collection for various biochemical and cell-based assays.

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Lab products found in correlation

3 3 5 5 tetramethylbenzidine (tmb), by Avantor (1 mentions) 96 well plate, by Greiner (1 mentions) Thrombin, by Siemens (1 mentions) Adenosine diphosphate (adp), by Merck Group (1 mentions) Lipopolysaccharides (lps), by Merck Group (1 mentions) Vacutainer sst 2 advance, by BD (1 mentions) Cell counting kit 8 (cck8), by Dojindo Laboratories (1 mentions) Annexin 5 fitc, by BD (1 mentions) Facscan flow cytometer, by BD (1 mentions)

Close spelling variants of this product

ChroMate microplate reader Chromate 4300 Microplate reader ChroMate

8 protocols using chromate 4300 microplate reader

ELISA Protocol for SARS-CoV-2 RBD Antibody Detection

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As an antigen for the enzyme-linked immunosorbent assay (ELISA), the RBD protein produced in CHO-K1 cells and purified using affinity and ion-exchange chromatography methods (protein purity > 98%) was used [30 (link)]. RBD protein (1 µg/mL in 2 m urea) was adsorbed onto 96-well plates (Greiner Bio-One, Kremsmünster, Austria) at 4 °C overnight, then the plate was washed in PBS with 0.05% Tween 20 (PBST) and blocked with 1% casein solution in the same buffer for 60 min at room temperature. Then, sera were added to the wells in threefold serial dilutions, starting from 1:50, in blocking solution, and incubated for 60 min at room temperature. The plate was washed, and horseradish-peroxidase-conjugated rabbit anti-mouse IgG antibodies (Sigma, St. Louis, MO, USA) were added and incubated for 60 min at room temperature. The substrate 3,3’,5,5’-tetramethylbenzidine (TMB) (Amresco, Dallas, TX, USA) was introduced into the wells of the plate washed with PBST. Optical density was measured at a wavelength of 450 nm using a ChroMate-4300 microplate reader (Awareness Technology Inc., Palm City, FL, USA). Serum dilution was taken as the titer in ELISA, at which the optical density value was more than two times higher than that for the negative control (blocking buffer was added to the wells instead of serum).

Kisakov D.N., Kisakova L.A., Borgoyakova M.B., Starostina E.V., Taranov O.S., Ivleva E.K., Pyankov O.V., Zaykovskaya A.V., Shcherbakov D.N., Rudometov A.P., Rudometova N.B., Volkova N.V., Gureev V.N., Ilyichev A.A, & Karpenko L.I. (2022). Optimization of In Vivo Electroporation Conditions and Delivery of DNA Vaccine Encoding SARS-CoV-2 RBD Using the Determined Protocol. Pharmaceutics, 14(11), 2259.

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Evaluating Antimicrobial Peptides' Activity

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The antibacterial activity of peptides was evaluated using a standard broth microdilution protocol ⁶⁶ with minor modifications as described.^{47 (link)} In brief, peptides (10 μL per well) with two fold dilutions were made in 96-well polystyrene microplates. Subsequently, the logarithmic phase bacterial cultures (i.e., optical density at 600 nm ≈ 0.5) were diluted to 0.001 OD and aliquoted 90 μL per well. The plates were incubated overnight at 37 °C. Bacterial controls were treated with water and the wells containing only the TSB media were used as blank. Positive controls include parent peptide LL-37 as well as KR12 discovered by our lab.^{33 (link)} Post incubation the plates were read at 630 nm using ChroMate4300 Microplate Reader (Awareness Technology, FL). The minimal inhibitory concentration (MIC) was the lowest peptide concentration that fully inhibited bacterial growth.
To study the influence of medium conditions on the antimicrobial activity of peptides against S. aureus USA300, different, pH, sodium chloride (NaCl), human sera were included into the antibacterial assays. Control experiments in TSB without any additions (pH 7.4) were set up in the same manner. In addition, antimicrobial activities of human LL-37, KR12, and C10-KR8d were compared in 10%, 50%, and 100% TSB as well as in TSB and MHB using E. coli ATCC 25922 and S. aureus USA300 LAC.

Narayana J.L., Golla R., Mishra B., Wang X., Lushnikova T., Zhang Y., Verma A., Kumar V., Xie J, & Wang G. (2021). Short and Robust Anti-infective Lipopeptides Engineered Based on the Minimal Antimicrobial Peptide KR12 of Human Cathelicidin LL-37. ACS infectious diseases, 7(6), 1795-1808.

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Quantification of Plasma Alarin and Adipsin

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Plasma Alarin and Adipsin levels were examined using the Human Alarin, Adipsin ELISA Kit (Sunred Biological Technology, Shanghai, China) in a plate-washing -incubation CombiWash device (Human Diagnostics, Wiesbaden, Germany) in accordance with the study procedures determined in the kit catalogue, and the absorbance measurement was taken with a Chromate 4300 Microplate Reader (Awareness Technology, Palm City, USA).
Aqueous analyzes were performed according to a previously published methods [15 (link)]. Two Aqueous liquids and blood samples were enriched with increasing amounts of Adipsin or Alarin. The percentage recovery was calculated as follows: recovered value/expected value × 100.
The measurement range human alarin kit was 5 to 1500 pg/mL and the sensitivity was determined by the manufacturer at 4.638 pg/mL. The intra-assay and inter-assay coefficients of variation for alarin were < 10% and < 12%, respectively. The measurement range of the human Adipsin kit 0.5 to 100 ng/mL and the sensitivity was the determined by the manufacturer at 0.472 ng/mL. The intra-assay and inter-assay coefficients of variation for Alarin were < 10% and < 12%, respectively.

Gül F.C., Kobat S.G., Çelik F., Aydin S, & Akkoç R.F. (2022). Plasma and aqueous levels of alarin and adipsin ın patients with and without diabetic retinopathy. BMC Ophthalmology, 22, 176.

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Azurocidin Secretion from Stimulated Cells

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Meg-01 cells (1 × 10⁶) and platelets (1 × 10⁷) were collected and stimulated with 1 unit of thrombin (Dade Behring, Deerfield, IL, USA), 5 μM of ADP (Sigma-Aldrich), or 1 μg of LPS (Sigma-Aldrich), and apyrase was used to benefit its baseline condition of non-activation. (0.25 U/mL) (Sigma-Aldrich) was used, and all stimuli were incubated at 37 °C for 30 min. The supernatant was collected, and the secreted azurocidin was quantified by ELISA using an optical density. Readings were performed at 450 nm using the Chromate 4300 microplate reader (Awareness Technology Inc.; Palm City, FL, USA) with the Leica Suite 345 X software application (LAS-X, Leica-Microsystems).

Aquino-Domínguez A.S., Acevedo-Sánchez V., Cruz-Hernández D.S., Sánchez-Aparicio S.R., Romero-Tlalolini M.D., Baltiérrez-Hoyos R., Sánchez-Navarro L.M., Torres-Aguilar H., Bustos-Arriaga J, & Aguilar-Ruiz S.R. (2022). Human Platelets Contain, Translate, and Secrete Azurocidin; A Novel Effect on Hemostasis. International Journal of Molecular Sciences, 23(10), 5667.

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Determining Antimicrobial Peptide Resistance

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This experiment performed similar to the MIC determination with a few modifications as described.^{47 (link)} In short, an exponential phase S. aureus USA300 culture (i.e., optical density at 600 nm ≈ 0.5) was diluted and partitioned into a 96-well polystyrene microplate with ~10⁵ bacteria per well (90 μL aliquots). After treatment with 10 μL of peptide or nafcillin solutions at various concentrations, microplates were incubated at 37 °C overnight and read on a ChroMate 4300 Microplate Reader at 630 nm (Awareness Technology, FL, USA). The wells with sub-MIC levels of the peptides that retained growth approximately half the growth of the control wells were again re-inoculated in fresh TSB with sub-MIC concertation of peptides or antibiotics to attain exponential phase for MIC determination. Up to 15 serial passages of the bacteria cultures were conducted. The increase in the fold change (MIC on given passage/ MIC recorded in first day of passage) used to determine the degree of drug resistance.

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Plasma Peptide Levels Measurement

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Plasma subfatin, preptin, and betatrophin levels were examined using the Human subfatin, preptin, betatrophin ELISA Kit (Sunred Biological Technology, Shanghai, China) in a plate-washing -incubation CombiWash device (Human Diagnostics, Wiesbaden, Germany) in accordance with the study procedures defined in the kit catalogue. Absorbance measurements were taken with a Chromate 4300 Microplate Reader (Awareness Technology, Palm City, USA).
These kit companies in their kit catalogs stated that the minimum detection limit of subfatin was 0.042 ng/mL. The intra-assay and inter-assay coefficients of variation for plasma subfatin were < 10% and < 12%, respectively. The minimum detection limit of preptin was 5.125 ng/L (0.0051 ng/mL). The intra-assay and inter-assay coefficients of variation for plasma preptin were < 10% and < 12%, respectively. The minimum detection limit of betatrophin was 7.334 ng/L (0.0073 ng/mL). The intra-assay and inter-assay coefficients of variation for plasma betatrophin were < 10% and < 15%, respectively. Preptin and betatrophin concentration units are given in ng/L and subfatin in ng/mL by the manufacturer. However, to facilitate the comparison between parameters, all units were converted to ng/mL [20 (link)].

Güngör Kobat S., Gül F.C., Çelik F., Liman Uzun S., Kobat M.A., Akkoç R.F, & Aydın S. (2023). Plasma and aqueous levels of subfatin, preptin and betatrophin in patients with diabetic retinopathy. BMC Ophthalmology, 23, 312.

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Plasma Metallothionein-1 Quantification

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In the study, samples were taken from the control and patient groups after 12 h of fasting into a tube containing aprotinin (BD Vacutainer SST II Advance, BD, Plymouth, UK). Blood samples were centrifuged at 2000–3000 rpm for 20 min and plasma containing Metallothionein-1 was placed in small volume tubes for analysis and stored at −20 °C until the study day.
Plasma Metallothionein-1 levels were studied using the Human Metallothionein-1 ELISA Kit (Bioassay Technology Laboratory, catalog no: E4358hu, Shanghai, China) in accordance with the study procedures specified in the kit catalog; absorbance measurement was made using a Chromate 4300 Microplate Reader (Awareness Technology, Palm City, FL, USA) device. The minimum detection limit of Metallothionein-1 was 0.24 ng/mL. The intra-assay and inter-assay coefficient of variation for plasma Metallothionein-1 were <8% and <10%, respectively.

Yılmaz S., Kılıç N., Kaya Ş, & Taşcı G. (2023). A Potential Biomarker for Predicting Schizophrenia: Metallothionein-1. Biomedicines, 11(2), 590.

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Cell Viability and Apoptosis Assays

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The cell viability test was conducted by the Cell Counting Kit‐8 (CCK‐8, Dojindo, Kumamoto, Japan) based on the quantity of a formazan dye, referring to the suggestion of producers. The ChroMate 4300 microplate reader (Awareness Technology, Palm City, FL, USA) was used to evaluate the absorbance set at OD 450 nm. The flow cytometry for cell apoptosis was carried out using fluorescein isothiocyanate labeled Annexin V (Annexin V‐FITC, BD Biosciences, Cowley, UK) and propidium iodide (PI, BD Biosciences) staining as described previously.17 A FACScan flow cytometer (BD Biosciences) was employed to detect and quantify apoptotic cells.

Liu G., Wan Q., Li J., Hu X., Gu X, & Xu S. (2020). Circ_0038467 regulates lipopolysaccharide‐induced inflammatory injury in human bronchial epithelial cells through sponging miR‐338‐3p. Thoracic Cancer, 11(5), 1297-1308.

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