Methods of performing a sizing analysis using a corrected sizing ladder
Inventors
CAMPORESE, DAN • Bhatt, Advit • Molho, Josh • Xu, Hui • Summers, Ken
Assignees
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Publication Number
US-8871524-B2
Publication Date
2014-10-28
Expiration Date
Abstract
The invention provides methods of performing a sizing analysis. In the methods, a sizing ladder used in performing the sizing analysis is corrected. In one method, the sizing ladder is corrected for batch-to-batch variations in a sieving gel. In another method, the sizing ladder is corrected for a sample concentration that is different from the archival sizing ladder concentration. Methods are also provided in which the sizing ladder is corrected using a standard marker in a sample and/or using a real-time standard sizing ladder. The methods may be used individually or in combination.
Core Innovation
The invention provides a method of performing a sizing analysis of separated components by using an archival sizing ladder that includes previously measured archival ladder migration times. Separated components from a sample are analyzed in a fractionation device using a corrected archival sizing ladder, and the corrected ladder is derived from migration times measured for a standard sizing ladder through a separation channel that contains a quantity of a batch of sieving gel.
To address sieving-gel batch-to-batch variation, the method measures migration times for a standard sizing ladder to provide a sieving gel sizing ladder with sieving gel ladder migration times. A ratio is determined between the sieving gel ladder migration times and the archival ladder migration times, thereby providing a first correction, and the first correction is encoded into first information storage means, attached to a second fractionation device, and then read and applied to each archival ladder migration time to provide a first corrected archival sizing ladder.
Sizing analysis of the separated components is then performed by comparing the sample to the first corrected archival sizing ladder using a parameter selected from separated component migration time and separated component band position. Additional corrections are described that update an archival ladder using real-time standard ladder migration information and that correct for differences in sample concentration versus ladder concentration by normalizing cumulative sample concentration and applying an empirical concentration-based correction to ladder peak migration times.
Claims Coverage
The document provides one independent claim covering a complete workflow for generating and propagating a corrected archival sizing ladder using batch-specific sieving gel measurements, encoding the correction for use in another fractionation device, and performing sizing analysis using migration time and/or band position. The claim set includes dependent refinements that constrain the magnitude of corrections and specify the form of the information storage, and further extend correction updates across additional fractionation devices and sieving-gel batches.
Archival sizing ladder with previously measured migration times
Providing an archival sizing ladder, the archival sizing ladder including previously measured archival ladder migration times.
Batch-specific calibration using a sieving-gel sizing ladder
Measuring migration times for a standard sizing ladder through a separation channel in a first fractionation device, the separation channel having a quantity of a first batch of sieving gel disposed therein, thereby providing a first sieving gel sizing ladder, the first sieving gel sizing ladder including first sieving gel ladder migration times.
Ratio-based correction between sieving-gel and archival migration times
Determining a ratio of the first sieving gel sizing ladder migration times to the archival ladder migration times, thereby providing a first correction.
Encoded correction carried in information storage attached to another fractionation device
Encoding the first correction into a first information storage means; attaching the first information storage means to a second fractionation device, a separation channel of the second fractionation device having a quantity of the first batch of sieving gel disposed therein; reading the first correction encoded into the information storage means.
Applying encoded correction to archival ladder to create a corrected archival sizing ladder
Applying the first correction to each archival ladder migration time, thereby providing a first corrected archival sizing ladder.
Sizing analysis using corrected ladder with migration time or band position
Performing a sizing analysis of the separated components of a sample disposed in the second fractionation device using the first corrected archival sizing ladder in comparison with a parameter for at least one of the separated components, the parameter being selected from the group consisting of separated component migration time and separated component band position.
Overall, the claim coverage centers on generating a first correction by ratioing sieving-gel ladder migration times against previously measured archival ladder migration times, encoding that correction in information storage carried to another fractionation device, applying the correction to produce a corrected archival sizing ladder, and using that ladder to perform sizing analysis using separated component migration time and/or band position. Dependent claims further specify quantitative correction constraints and an information-storage form, and extend correction updates using additional batches and additional fractionation devices.
Stated Advantages
Improves sizing analysis accuracy by correcting for batch-to-batch sieving-gel variation using measured standard sizing ladder migration times.
Enables continued, device-to-device use of corrected sizing based on an encoded correction that is read and applied in a second fractionation device.
Supports real-time/updated sizing by updating archival ladder information using detected real-time standard ladder information.
Improves sizing accuracy by correcting for differences between sample concentration and ladder concentration using a concentration-relative correction.
Documented Applications
Sizing analysis of separated components by comparing separated component migration time and/or separated component band position to a corrected archival sizing ladder in a fractionation device.
Real-time fractionation/diversion timing accuracy using corrected or updated sizing ladder migration information.
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