2007
DOI: 10.1002/cyto.a.20507
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Toward quantitative fluorescence measurements with multicolor flow cytometry

Abstract: A procedure is presented for calibrating the output of a multicolor flow cytometer in units of antibodies bound per cell (ABC). The procedure involves two steps. First, each of the fluorescence channels of the flow cytometer is calibrated using Ultra Rainbow beads with assigned values of equivalent number of reference fluorophores (ERF). The objective of this step is to establish a linear relation between the fluorescence signal in a given fluorescence channel of multicolor flow cytometers and the value of ERF… Show more

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Cited by 66 publications
(70 citation statements)
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“…To distinguish this approach from MESF, the fluorescence intensity unit used for assignment is called equivalent number of reference fluorophores (ERF) (2). As long as this is restricted to a particular instrument model, the data in this report indicates the approach can be used if variation of 10% or more is acceptable.…”
Section: Discussionmentioning
confidence: 99%
“…To distinguish this approach from MESF, the fluorescence intensity unit used for assignment is called equivalent number of reference fluorophores (ERF) (2). As long as this is restricted to a particular instrument model, the data in this report indicates the approach can be used if variation of 10% or more is acceptable.…”
Section: Discussionmentioning
confidence: 99%
“…An advantage of using the %CV rather than the SD is that it normalizes variations at lower levels of event detection. Considerations when establishing the level of acceptable imprecision for a reportable result include the frequency of the population and the total number of events acquired (11,12). A desirable target for assay imprecision is a CV of less than 10%, but for less abundant populations (where frequency is at a level of 1:1,000 (0.1%) or lower), such as minimal residual disease detection or fetomaternal hemorrhage detection, a CV of less than 20% may be acceptable (13).…”
Section: Intra-assay Imprecisionmentioning
confidence: 99%
“…For example, the linearity of population frequency may be estimated by assay of a positive sample prepared by known serial dilution into a negative sample, as above. Only when the fluorescence intensity signal output is quantified using fluorescence calibration/quantitation beads is the assay considered relatively quantitative, in which case linearity should be evaluated using alternative approaches to serial dilutions (11). On the other hand, instrument linearity can be demonstrated and should be verified semi-annually per the laboratory's instrument SOP; this is sufficient and appropriate assessment of linearity for relative quantitative assays.…”
Section: Linearitymentioning
confidence: 99%
“…Additionally, binding of these aptamers to cell surface proteins is proposed to interfere with the function of the protein and may result in different phenotypes and potentially negatively influence the parasite. Moreover, together with trafficking analysis of GFP-labeled P. falciparum chimer proteins, aptamers identified binding to distinct parasite proteins on infected erythrocytes can be derivatized by quantum dots and used for multiparameter cytomics (73,74).…”
Section: Employing Aptamers On Parasite Infectionmentioning
confidence: 99%