Plate reader: Difference between revisions

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{{more citations needed|date=April 2012}}
[[File:Microplate reader.jpg|thumb|[[BioTek]] PowerWave XS Microplate Reader]]
 
'''Plate readers''', also known as '''microplate readers''' or '''microplate photometers''', are instruments which are used to detect [[biology|biological]], [[chemistry|chemical]] or [[physics|physical]] events of samples in [[microtiter plate]]s. They are widely used in research, [[drug discovery]],<ref>{{Citecite journal |last1=Neves |first1=Bruno Junior |last2=Agnes |first2=Jonathan Paulo |last3=Gomes |first3=Marcelo do Nascimento |last4=Henriques Donza |first4=Marcio Roberto |last5=Gonçalves |first5=Rosângela Mayer |last6=Delgobo |first6=Marina |last7=Ribeiro de Souza Neto |first7=Lauro |last8=Senger |first8=Mario Roberto |last9=Silva-Junior |first9=Floriano Paes |last10=Ferreira |first10=Sabrina Baptista |last11=Zanotto-Filho |first11=Alfeu |datelast12=2020-03-01Andrade |first12=Carolina Horta |title=Efficient identification of novel anti-glioma lead compounds by machine learning models|url=http://www.sciencedirect.com/science/article/pii/S022352341931133X |journal=European Journal of Medicinal Chemistry |languagedate=enMarch 2020 |volume=189 |pages=111981 |doi=10.1016/j.ejmech.2019.111981 |pmid=31978780 |s2cid=210892159 |issn=0223-5234}}</ref> bioassay validation, quality control and manufacturing processes in the pharmaceutical and biotechnological industry and academic organizations. Sample reactions can be assayed in 1-1536 well format microtiter plates. The most common microplate format used in academic research laboratories or clinical diagnostic laboratories is 96-well (8 by 12 matrix) with a typical reaction volume between 100 and 200 µLμL per well. Higher density microplates (384- or 1536-well microplates) are typically used for screening applications, when throughput (number of samples per day processed) and assay cost per sample become critical parameters, with a typical assay volume between 5 and 50 µLμL per well. Common detection modes for microplate assays are absorbance, [[fluorescence]] intensity, [[luminescence]], [[Time-resolved spectroscopy#Time-resolved fluorescence spectroscopy|time-resolved fluorescence]], and [[fluorescence polarization]].
 
==Methods==
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===Fluorescence===
Fluorescence intensity detection has developed very broadly in the microplate format over the last two decades. The range of applications is much broader than when using absorbance detection, but the instrumentation is usually more expensive. In this type of instrumentation, a first optical system (excitation system) illuminates the sample using a specific wavelength (selected by an optical filter, or a monochromator). As a result of the illumination, the sample emits light (it fluoresces) and a second optical system (emission system) collects the emitted light, separates it from the excitation light (using a filter or monochromator system), and measures the signal using a light detector such as a [[photomultiplier]] tube (PMT). The advantages of fluorescence detection over absorbance detection are sensitivity, as well as application range, given the wide selection of fluorescent labels available today. For example, a technique known as [[calcium imaging]] measures the fluorescence intensity of [[calcium-sensitive dyes]] to assess intracellular calcium levels.{{citation needed|date=May 2020}}<ref>{{Citecite journal |last1=Lin |first1=Kedan |last2=Sadée |first2=Wolfgang |last3=Mark Quillan |first3=J.|date=February 1999|title=Rapid Measurements of Intracellular Calcium Using a Fluorescence Plate Reader |journal=BioTechniques |languagedate=enFebruary 1999 |volume=26 |issue=2 |pages=318–326 |doi=10.2144/99262rr02 |pmid=10023544 |issn=0736-6205|doi-access=free }}</ref>
 
===Luminescence===
Luminescence is the result of a chemical or biochemical reaction. Luminescence detection is simpler optically than fluorescence detection because luminescence does not require a light source for excitation or optics for selecting discrete excitation wavelengths. A typical luminescence optical system consists of a light-tight reading chamber and a [[Photomultiplier|PMT]] detector. Some plate readers use an Analog PMT detector while others have a [[photon counting]] PMT detector. Photon Counting is widely accepted as the most sensitive means of detecting luminescence. Some plate readers offer filter wheel or tunable wavelength monochromator optical systems for selecting specific luminescent wavelengths. The ability to select multiple wavelengths, or even wavelength ranges, allows for detection of assays that contain multiple luminescent reporter enzymes, the development of new luminescence assays, as well as a means to optimize the signal to noise ratio.{{citation needed|date=May 2020}}
 
Common applications include [[luciferase]] -based gene expression assays, as well as cell viability, cytotoxicity, and biorhythm assays based on the luminescent detection of [[Adenosine triphosphate|ATP]].<ref>{{Citecite journal |last1=Lin |first1=Kedan |last2=Sadée |first2=Wolfgang |last3=Mark Quillan |first3=J.|date=1999-02-01 |title=Rapid Measurements of Intracellular Calcium Using a Fluorescence Plate Reader |journal=BioTechniques |date=February 1999 |volume=26 |issue=2 |pages=318–326 |doi=10.2144/99262rr02 |pmid=10023544 |issn=0736-6205|doi-access=free }}</ref>
 
===Time-resolved fluorescence (TRF)===
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*[[ELISA]]s
*Protein and [[cell growth]] assays
*[[Protein–protein interaction]]
*Protein:protein interactions
*[[Reporter gene|Reporter]] assays
*[[Nucleic acid quantitation]]
*Molecular interactions
*Enzyme activity
*Cell toxicity, proliferation, and viability
*[[ATP test|ATP quantification]]
*[[Immunoassays]]<ref>{{Citecite journal |last1=Ashour |first1=Mohamed-Bassem A. |last2=Gee |first2=Shirley J. |last3=Hammock |first3=Bruce D.|date=1987-11-01 |title=Use of a 96-well microplate reader for measuring routine enzyme activities|url=https://dx.doi.org/10.1016/0003-2697%2887%2990585-9 |journal=Analytical Biochemistry |languagedate=enNovember 1987 |volume=166 |issue=2 |pages=353–360 |doi=10.1016/0003-2697(87)90585-9 |pmid=3434778 |issn=0003-2697}}</ref>
*[[High throughput screening]] of compounds and targets in drug discovery (LabelLabeled Alpha Screen on most instruments)<ref>{{cite web | url=https://www.bmglabtech.com/en/alphascreen/ | title=AlphaScreen &#124; BMG LABTECH }}</ref>
*Bead-Basedbased Epitope[[epitope]] Assayassay<ref>{{cite journal |last1=Suprun |first1=Maria |last2=Getts |first2=Robert |last3=Raghunathan |first3=Rohit |last4=Grishina |first4=Galina |last5=Witmer |first5=Marc |last6=Gimenez |first6=Gustavo |last7=Sampson |first7=Hugh A. |last8=Suárez-Fariñas |first8=Mayte |title=Novel Bead-Based Epitope Assay is a sensitive and reliable tool for profiling epitope-specific antibody repertoire in food allergy |journal=Scientific Reports |date=5 December 2019 |volume=9 |issue=1 |page=18425 |doi=10.1038/s41598-019-54868-7 |pmid=31804555 |pmc=6895130 |bibcode=2019NatSR...918425S }}</ref>
*Cellular Uptake of [[nanoparticle]]s<ref>{{cite journal | doi=10.1038/s41598-022-24480-3 | title=Quantifying fluorescent nanoparticle uptake in mammalian cells using a plate reader | year=2022 | last1=Shin | first1=Hye Ji | last2=Kwak | first2=Minjeong | last3=Joo | first3=Sihwa | last4=Lee | first4=Ji Youn | journal=Scientific Reports | volume=12 | issue=1 | page=20146 | pmid=36418509 | pmc=9684140 | bibcode=2022NatSR..1220146S }}</ref>
*Cellular Uptake of nanoparticles<ref>https://www.nature.com/articles/s41598-022-24480-3</ref>
 
While "plate reader" usually refers to the devices described above, many variations are available. Some examples of other devices working with the microplate format are: