On Wed, 10 May 2000, Bennett, Catherine (Katie) wrote:

> Got a co-worker who's not on the net looking for information on Annexin V
> labeling/staining for apoptosis analysis in rat lung tissues.  Has anyone
> out there done this before in tissue? 

  Annexin V binds specifically to phosphatidylserine, a lipid 
  that is normally on the inside of the cell membrane but which
  is shifted to the outside surface of the cell quite early in
  the apoptotic process. Labelled annexin V therefore sticks to
  whole cells (with intact membranes) that are undergoing apoptosis,
  but not to healthy cells. (Annexin V is a protein, so it can't
  get through the intact cell membrane.)

  If you were to apply labelled annexin V to a section, it would
  bind to phosphatidylserine on the inside surfaces of normal cell
  membranes as well as to the outsides of apoptotic cells. With 
  light microscopy of any kind the resolution is far too low to
  distinguish one side of a membrane (thickness about 7.5nm) from
  the other. The book "Techniques in Apoptosis" ed. TG Cotter &
  SJ Martin; London: Portland Press (1996) has a chapter with
  explanations and instructions for using annexin V (pp 107-119).

  There have been several published comparisons of different
  methods for detection of apoptotic cells, with electron
  microscopy (TEM) as the absolute standard. Generally, the
  methods for fragmented DNA (TUNEL and in situ nick 
  translation) detected fewer apoptotic cells than simple 
  nuclear staining with either alum-haematoxylin or a stain 
  for DNA such as a simple basic dye or fluorochrome. The 
  book mentioned in the previous paragraph has som nice colour 
  photos of apoptotic bodies in the intestine, stained with
  H & E (on page 283).
 
 John A. Kiernan,
 Department of Anatomy & Cell Biology,
 The University of Western Ontario,
 LONDON,  Canada  N6A 5C1