[Histonet] Re: lymphatics

From:"Andrea T. Hooper"

Hi Histonet,

Has anyone performed the 5-nucleotidase stains John referred to 
below. I am really interested in this method of distinguising BV from 
LV. If so does someone have a working protocol I can work from or 
words of wisdom??

Thanks in advance,

>Dear Hal Hawkins,
>The endothelium of lymphatic vessels can be stained by
>enzyme activity histochemistry. The enzyme is a
>5-nucleotidase (belongs to the phosphatase family).
>Ordinarily this type of method is carried out with frozen
>sections of tissue that has been minimally fixed in
>formaldehyde or glutaraldehyde, or with unfixed cryostat
>sections that have been briefly fixed in cold acetone. The
>enzyme in lymphatics may be fairly robust because its
>activity can survive embedding in glycol methacrylate.
>The references below may help. If you choose one of these
>methods it will be important to consult the original
>publication because there's more than one sort of
>5-nucleotidase. Each ref is followed by brief notes that I
>took when reading the papers. [An R in the acquisition
>number means I've got a Reprint of the whole paper; an A
>usually means I have a copy of the paper's Abstract. No
>letter with the acquisition number usually means I've seen
>the whole paper and took notes in the library.]
>8194R. Kato,S; Yasunaga,A; Uchida,U (1991):
>Enzyme-histochemical method for identification of lymphatic
>capillaries. Lymphology 24, 125-129.
>   Glycol methacrylate-embedded sections stained for
>5'-nucleotidase & alkaline phosphatase. 5-N-ase in lymph
>capills; AlkP-ase in blood capills.
>9239R. Nishida,S; Ohkuma,M (1993): Enzyme-histochemical
>staining of dermal lymphatic capillaries by guanylate
>cyclase. Lymphology 26, 195-199.
>   Enzyme histochemical staining distinguishes lymph from
>blood capillaries. Says 5-nucleotidase & adenylate cyclase
>do so too. (Used unfixed cryostat sections.)
>9665R. Okada,E (1994): An improved enzyme-histochemical
>method for identification of lymphatic capillaries on
>paraffin sections. Lymphology 27, Suppl, 732-735.
>   Staining of blood & lymph capillaries. Enzyme
>histochemistry for 5-nucleotidase in presence of levamisole
>for lymph capills; alkaline phosphatase for blood capills.
>11114A. Miura,M; Kato,S; von Ludinghausen,M (1998):
>Lymphatic drainage of the cerebrospinal fluid from monkey
>spinal meninges with special reference to the distribution
>of the epidural lymphatics. Arch. Histol. Cytol. 61(3, Aug),
>    5'-nucleotidase staining for lymphatics; alk phosphatase
>for blood capillaries (Kato et al '91,'93). Also traced
>carbon particles from cisterna magna. Lymphatics and carbon
>found on surfaces of cervical & thoracic (most at brachial
>plexus levels) roots; not lumbosacral. Epidural lymphatics
>most developed on dorsal surface of lower cervical dura.
>I hope this helps.
John A. Kiernan

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