Re: [Histonet] staining for iron in microglia

From:John Kiernan

Sharon,  Your question is 100% scientific, 
not semi-scientific! You concisely provide an 
introduction, expand it into a testable hypothesis,
and ask if the test has been done. 

The answer is "Yes, but ..." 

Iron is seen in phagocytic cells in various pathological
conditions of the brain. Its presence in the frontal
cortex (seen with the regular Perls method) is a 
classical feature of advanced syphilis (general paralysis
of the insane). There are some cells in the normal rat's
hypothalamus that have a variety of histochemical
peculiarities including being Perls-positive, but these 
cells are astrocytes (Young & McKenzie, 2004).

The DAB-enhanced Perls method shows non-haem iron in 
microglia, oligodendroglia and neurons. Research in this 
field, from several labs, was published in the late 1980s 
and early 1990s, about 10 years after the publication of 
the DAB amplification procedure by
Nguyen-Legros et al in 1980. A few references are given
at the end of this message. They are ones that I've seen
and made notes on; there will be many more, findable by
way of PubMed, Web of Science or Scopus. At NIH all
those resources and more will be available to you. 

Another iron-related approach to microglia has been
immunostaining for ferritin (Kaneko et al 1989). There
are some lectins with affinity for microglia that can
provide impressive pictures when correctly applied.
For the old and bold there are the original silver 
stains, developed by Del Rio Hortega and perfected by
Penfield and Cone in the 1920s. 

Summary.  Sensitive methods for iron do not provide
specific staining of resident, activated or immigrant
microglial cells. The classical Perls method can 
detect microglia that have collected iron in sites of
injury or disease. Iron is also present in macroglia 
and neurons, and is stainable in these cells if the 
sensitivity of the Perls prussian blue method is 


Gerber MR, Connor JR (1989) Do oligodendrocytes mediate iron
regulation in the human brain? Ann. Neurol. 26: 95-98.

Kaneko Y, Kitamoto T, Tateishi J, Yamaguchi K (1989) Ferritin
immunohistochemistry as a marker for microglia. Acta Neuropathol.
(Berl.) 79: 129-136.

Morris CM, Candy JM, Oakley AE, Bloxham CA, Edwardson JA (1992)
Histochemical distribution of non-haem iron in the human brain.
Acta Anat. 144: 235-257.

Connor JR, Pavlick G, Karli D, Menzies SL, Palmer C (1995) A
histochemical study of iron-positive cells in the developing rat
brain. J. Comp. Neurol. 355: 111-123.

Palmer C, Menzies SL, Roberts RL, Pavlick G, Connor JR (1999)
Changes in iron histochemistry after hypoxic-ischemic brain
injury in the neonatal rat. J. Neurosci. Res. 56: 60-71.

Young JK, McKenzie JC (2004) GLUT2 immunoreactivity in
Gomori-positive astrocytes of the hypothalamus. J. Histochem.
Cytochem. 52: 1519-1524.

John Kiernan
Anatomy, UWO
London, Canada
Sharon Cooperman wrote:
> Dear Histonetters,
> I have a semi-scientific question:  macrophages in almost all tissues
> can be stained for iron with Perl's stain to give a blue color - is
> this also true for microglia?  If not, is there any other way to see
> how much iron is in microglia (DAB enhanced Perl's or some other
> method)?
> Thanks,
> Sharon
> --
> Sharon Cooperman                     
> NIH, NICHD, CBMB                     301.435-8417
> Building 18T, room 101               301.402-0078 fax
> Bethesda, MD 20892
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