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#solute elution

3 public questions tagged with this topic.

Chromatography

Elute first in size exclusion chromatography?

Size exclusion chromatography separates strictly by hydrodynamic volume, not by charge or affinity. Porous inert beads allow small molecules to diffuse into pores, increasing path length and retention, while large molecules unable to enter are restricted to void space between beads. Consequently, large molecules travel shortest distance and emerge first, followed by progressively smaller species. This elution order opposite to many other methods enables molecular weight estimation, desalting, and removal of aggregates. Understanding inverse relationship between size and retention is crucial fo

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Chromatography

Elutes first in cation exchange at pH 7?

Cation exchange matrix bears negative functional groups attracting positively charged species. At pH 7, acidic amino acids like aspartate with pI around 3 are deprotonated and negatively charged, thus repelled and eluted first without binding. Basic residues like lysine remain positively charged and bind strongly. Neutral aliphatic residues such as valine and leucine carry little net charge at neutral pH, resulting in minimal interaction. Hence in a mixture, aspartate exits earliest, allowing charge-based fractionation. Adjusting pH relative to amino acid pI values facilitates predictable elut

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

View Details
Chromatography

Elute last in DEAE column:

DEAE, diethylaminoethyl cellulose, is a weak anion exchanger carrying positively charged groups that attract negatively charged analytes. Binding strength correlates with net negative charge density at given pH. Molecules with higher negative charge interact more strongly, requiring higher salt concentration for displacement. Therefore, among proteins, the most acidic, most negatively charged species retains longest and elutes last during ionic strength gradient, whereas positively charged or neutral proteins pass through or elute early. This principle enables separation of isoforms, nucleic a

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

View Details