Numerous laboratories are focusing efforts on delivering gene products to i
nduce or prevent the development of new blood vessels in adults, with the h
ope of rescuing ischemic tissues, circumventing cardiac bypass surgery, or
inhibiting tumor growth. Current approaches to the assessment of vascular c
ontinuity involve the introduction of either dyes or fluorescent microspher
es to track blood flow. However, dyes and dextrans are subject to leakage w
hen vessels are hyperpermeable, a situation that may occur in studies of tu
mor vasculature and during efforts to stimulate therapeutic angiogenesis. F
urthermore, the microspheres that are used for flow studies do not allow a
comprehensive visual analysis of vascular continuity. Here we report a meth
od for the visual assessment of microvascular continuity in mouse muscle un
der circumstances in which vessels are leaky. The approach involves perfusi
on of the vasculature with fluorescent beads that are much smaller than tho
se used for flow studies. The suspension behaves like a fluid and completel
y fills the vessels, yet the beads do not leak from VECF-permeablized capil
laries and remain localized in histological sections. Use of beads with the
proper fluorescence emission wavelengths allows immunofluorescent colocali
zation with vessel-specific markers. We compare this improved method with o
ther methods for tracking vascular continuity involving dextrans and larger
beads. This approach should aid in the dynamic study of tumor angiogenesis
and the evaluation of efforts to deliver angiogenic factors.