Correct Answer: A. P40 will run faster

In Sephadex G-100 gel filtration chromatography, smaller proteins (P40, 40 kDa) elute earlier (run faster) than larger proteins (P60, 60 kDa) because they penetrate more bead pores, taking a longer path through the column.

Option Analysis

A. P40 will run faster (Correct)
Gel filtration separates by hydrodynamic volume. P40 (smaller) enters more pores, travels longer internal path, elutes first. P60 (larger) excluded from pores, takes shorter external path, elutes later. Standard size-exclusion principle.

B. P60 will run faster
Incorrect. Larger proteins always elute earlier in gel filtration. P60 excluded from Sephadex G-100 pores (4-150 kDa range), cannot be true.

C. Always a 50% mixture of P40 and P60 will come out of the column
Incorrect. Complete baseline separation occurs between 40 kDa vs 60 kDa on G-100. No co-elution or fixed ratio expected.

D. P40 and P60 will come out in a ratio of 3:2 at any given time
Incorrect. Gel filtration yields discrete peaks by size, not concentration ratios. Elution timing independent of loading amounts.

P40 will run faster than P60 in Sephadex G-100 because smaller proteins elute first in gel filtration chromatography—fundamental principle for GATE Life Sciences protein purification.

Size Exclusion Mechanism

Sephadex G-100 (4-150 kDa range):

• P40 (40 kDa): Penetrates beads extensively, longer path → elutes first

• P60 (60 kDa): Partially excluded, shorter path → elutes second
  Linear log MW vs elution volume relationship ensures clean separation.

Elution Order Table

GATE Applications

Tests gel filtration fundamentals across biochemistry. Sephadex G-100 commonly used for 10-100 kDa protein desalting/purification in research protocols.