crosssectionatherosclerosis.pdf

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7/27/2019 CrosssectionAtherosclerosis.pdf http://slidepdf.com/reader/full/crosssectionatherosclerosispdf 1/3 Letters to the Editor 887 readily than microscopical examination of processed tissue,6 but this has not been systematically tested. These five cases of microsporidia represent 6 5% of the African HIV positive patients with intestinal symptoms that we saw. Three of the patients also had cryptosporidiosis (diagnosed on faecal smears, though not seen on histological examination). Irrespective of the degree of inflammation, no micro- sporidia have been identified in 36 small bowel biopsy specimens from HIV negative Ugandan and Zambian controls; no micro- sporidia have been seen in over 50 rectal biopsy specimens from HIV negative or HIV positive patients. More intestinal biopsy specimens from patients with AIDS should be examined for microsporidia. This group of parasites infects most phyla of invertebrates and all classes of vertebrates;yet E bieneusi has only been described in human enterocytes in association with HIV-1 infection.' It is unresolved whether they are genuinepatho- gens rather than mere passengers. The enterocytes containing microsporidia in our material and that of other workers do not show obvious damage by light or electron microscopy. The aetiology of the diarrhoea in AIDS is multifactorial, and microsporidia may be involved, possibly by affecting secre- tion. SB LUCAS* LUCIENNE PAPADAKI* C CONLONt N SEWANKAMBOt R GOODGAME D SERWADDA4 *Department of Histopathology, University College and Middlesex Hospital School of Medicine, University Street, London WCJ tDepartment of Medicine, University Teach- ing Hospital, Lusaka, Zambia, ,Department of Medicine, Mulago Hospital, Kampala, Uganda. References I Canning EU, Hollister WS. Microsporidia of mammals-widespread pathogens or oppor- tunistic curiosities? Parasitol Today 1987;3:267-72. 2 DesportesL, Le Charpentier Y, Galian A, et al. Occurrence of a new microsporidian: Enterocytozoon bieneusi n.g., n.sp., in the enterocytes of a human patient with AIDS. J Protozool 1985;32:250-4. 3 Dobbins WO, Weinstein W M. Electron micro- scopy of the intestine and rectum in acquired immunodefiency syndrome. Gastroenterol 1985;88:738-49. 4 Curry A, McWilliam LJ, Haboubi NY, Mandal BK. Microsporidiosis in a British patient with AIDS. J Clin Pathol 1988;41:477-8. 5 Modigliani R, Borles C, Le Charpentier Y, et al. Diarrhoea and malabsorption in acquired immunodeficiency syndrome: a study of four cases with special emphasis on opportunistic protozoal infestations. Gut 1985;26:179-87. 6 Rijpstra AC, Canning EU, van Ketel, RJ, et al. Use of light microscopy to diagnose small- intestinal microsporidiosis in patients with AIDS. JInfect Dis 1988;157:827-31. 7 Sewankambo N, Goodgame R, Carswell WJ, et al. Enteropathic AIDS in Uganda: a microbiological and histopathological study. AIDS 1987;1:9-13. 8 Nicholls G. Microsporidiosis in AIDS. Com- municable Diseases Report CDR 86/21. Lon- don: PHLS, 1986:3-4. 9 Brandborg LL, Goldberg SB, Breidenbach WC. Human coccidiosis-a possible cause of malabsorption. The life cycle in small-bowel biopsies as a diagnostic feature. N Engl J Me d 1970;283:1306-13. Visual aid for quick assessment of coronary artery stenosisat necropsy A chart was designed to give a quick visual assessment of the degree ofcoronary artery stenosis at necropsy. It was intended for routine cases in which coronary artery angiography and formal morphometric analysis of processed segments of vessel were not to be undertaken. Themethod described here is not the most accurate, but it is preferable to simple subjective categorisa- tion into simple, moderate, and severe degrees of artery stenosis. Ischaemic heart disease is one of the leading causes of death in Great Britain. Quantification of coronary artery stenosisas part of the necropsy is necessary to estimate the functional importance of any athero- sclerotic disease present. The cross sectional area of the vessel lumen, compared with that contained within the elastic lamina, expres- sed as a percentage, is a widely used method of estimating the degree of luminal area narrowing within a vessel.' A narrowing by 7 5% reduces coronary blood flow at times of stress and exertion; narrowing by 90% means that coronary blood flow is severely reduced at rest.2 Even in the absence of coronary thrombosis, cardiac death may occur if there is one segment of luminal area which has narrowed by more than 85%, although most of these patients have multi- ple foci of stenosis.3 Coronary angiography can be used routin- ely at necropsy4 to show the luminal diameter and it can also show all branches of the coronary vessels. Formal morphometry with the use of either point counting,5 or plan- imetry of multiple cross sections of vessels that have been fixed, decalcified, and stained for elastin, give a good estimation of luminal area narrowing. Vessels that have been per- fused and fixed at physiological pressures before dissection give the most meaningful results.6 An alternative, but the most routin- ely used method, is simple subjective assess- ment of a serially sliced vessel at the time of necropsy into categories of mild, moderate, and severe stenosis. The first two methods are obviously more accurate, and themerits and demerits of each have been discussed elsewhere.' The aim of our method is not to replace or compete with these methods but to make simple, immediate, subjective assessment more accurate and scientific. Examination ofcoronary arteries by serial transverse slicing at 3 mm intervals is widely practised. Most pathologists open the heart first and then slice the vessels while gripping the artery, with forefinger outside the heart and thumb inside. A sharp brain knife allows a "clean slice" to be made even with moderate vessel calcification. Once the vessels have been serially sliced, the cut surface is compared with the chart (figure). The vessels have been drawn with concentric round, eccentric round, and slit shaped lumina representing the three main patterns seen at necropsy. The diagrams have been photoreduced following the cal- culation of the areas on graph paper. Arteries that are distended at physiological pressure are almost circular in shape.' The (@) -90%-0 -90%- ( (i#I) -30%- K) -60- ((j) -70%- -70%- -60%- -60% - -40%- -40%- ______ -20% - ( -20%- Figure Schematic diagram representing a percentage reduction in cross sectional area of coronary artery vessels. The outer circle represents vesselexterior, inner circle represents the elastic lamina, and black area represents the lumen.  group.bmj.com on June 23, 2011 - Published by  jcp.bmj.com Downloaded from

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L e t t e r s t o t h e E d i t o r 8 8 7

r e a d i l y t h a n m i c r o s c o p i c a l e x a m i n a t i o n o f

p r o c e s s e d t i s s u e , 6 b u t t h i s h a s n o t b e e n

s y s t e m a t i c a l l y t e s t e d .T h e s e f i v e c a s e s o f m i c r o s p o r i d i a r e p r e s e n t

6 5% o f t h e A f r i c a n HIV p o s i t i v e p a t i e n t sw i t h i n t e s t i n a l s y m p t o m s t h a t w e s a w . T h r e e

o f t h e p a t i e n t s a l s o h a d c r y p t o s p o r i d i o s i s( d i a g n o s e d o n f a e c a l s m e a r s , t h o u g h n o t s e e n

o n h i s t o l o g i c a l e x a m i n a t i o n ) . I r r e s p e c t i v e o ft h e d e g r e e o f i n f l a m m a t i o n , n o m i c r o -s p o r i d i a h a v e b e e n i d e n t i f i e d i n 3 6 s m a l lb o w e l b i o p s y s p e c i m e n s f r o m HIV n e g a t i v eU g a n d a n a n d Z a m b i a n c o n t r o l s ; n o m i c r o -

s p o r i d i a h a v e b e e n s e e n i n o v e r 5 0 r e c t a lb i o p s y s p e c i m e n s f r o m HIV n e g a t i v e o r HIVp o s i t i v e p a t i e n t s .

M o r e i n t e s t i n a l b i o p s y s p e c i m e n s f r o m

p a t i e n t s w i t h AIDS s h o u l d b e e x a m i n e d f o rm i c r o s p o r i d i a . T h i s g r o u p o f p a r a s i t e si n f e c t s m o s t p h yl a o f i n v e r t e b r a t e s a n d a l lc l a s s e s o f v e r t e b r a t e s ; y e t E b i e n e u s i h a s o n l yb e e n d e s c r i b e d i n h u m a n e n t e r o c y t e s i n

a s s o c i a t i o n w i t h H I V - 1 i n f e c t i o n . ' I t i su n r e s o l v e d w h e t h e r t h e y a r e g e n u i n e p a t h o -

g e n s r a t h e r t h a n m e r e p a s s e n g e r s . T h ee n t e r o c y t e s c o n t a i n i n g m i c r o s p o r i d i a i n o u r

m a t e r i a l a n d t h a t o f o t h e r w o r k e r s d o n o t

s h o w o b v i o u s d a m a g e b y l i g h t o r e l e c t r o nm i c r o s c o p y . T h e a e t i o l o g y o f t h e d i a r r h o e ai n AIDS i s m u l t i f a c t o r i a l , a n d m i c r o s p o r i d i amay b e i n v o l v e d , p o s s i b l y b y a f f e c t i n g s e c r e -t i o n .

S B L U C A S *

L U C I E N N E P A P A D A K I *

C C O N L O N tN S E W A N K A M B O t

R G O O D G A M ED S E R W A D D A 4

* D e p a r t m e n t o f H i s t o p a t h o l o g y , U n i v e r s i t yC o l l e g e a n d M i d d l e s e x H o s p i t a l S c h o o l o f

M e d i c i n e , U n i v e r s i t y S t r e e t , L o n d o n W C Jt D e p a r t m e n t o f M e d i c i n e , U n i v e r s i t y T e a c h -

i n g H o s p i t a l , L u s a k a , Z a m b i a ,

, D e p a r t m e n t o f M e d i c i n e , M u l a g o H o s p i t a l ,K a m p a l a , U g a n d a .

R e f e r e n c e s

I C a n n i n g E U , H o l l i s t e r WS. M i c r o s p o r i d i a o f

m a m m a l s - w i d e s p r e a d p a t h o g e n s o r o p p o r -t u n i s t i c c u r i o s i t i e s ? P a r a s i t o l T o d a y

1 9 8 7 ; 3 : 2 6 7 - 7 2 .2 D e s p o r t e s L , L e C h a r p e n t i e r Y , G a l i a n A , e t a l .

O c c u r r e n c e o f a new m i c r o s p o r i d i a n :E n t e r o c y t o z o o n b i e n e u s i n . g . , n . s p . , i n t h ee n t e r o c y t e s o f a human p a t i e n t w i t h A I D S . J

P r o t o z o o l 1 9 8 5 ; 3 2 : 2 5 0 - 4 .3 D o b b i n s WO, W e i n s t e i n W M. E l e c t r o n m i c r o -

s c o p y o f t h e i n t e s t i n e a n d r e c t u m i n a c q u i r e di m m u n o d e f i e n c y s y n d r o m e . G a s t r o e n t e r o l1 9 8 5 ; 8 8 : 7 3 8 - 4 9 .

4 C u r r y A , M c W i l l i a m L J , H a b o u b i NY, M a n d a l

B K . M i c r o s p o r i d i o s i s i n a B r i t i s h p a t i e n t w i t h

A I D S . J C l i n P a t h o l 1 9 8 8 ; 4 1 : 4 7 7 - 8 .

5 M o d i g l i a n i R , B o r l e s C , L e C h a r p e n t i e r Y , e t a l .

D i a r r h o e a a n d m a l a b s o r p t i o n i n a c q u i r e di m m u n o d e f i c i e n c y s y n d r o m e : a s t u d y o f f o u rc a s e s w i t h s p e c i a l e m p h a s i s o n o p p o r t u n i s t i cp r o t o z o a l i n f e s t a t i o n s . G u t 1 9 8 5 ; 2 6 : 1 7 9 - 8 7 .

6 R i j p s t r a A C , C a n n i n g E U , v a n K e t e l , R J , e t a l .

U s e o f l i g h t m i c r o s c o p y t o d i a g n o s e s m a l l -i n t e s t i n a l m i c r o s p o r i d i o s i s i n p a t i e n t s w i t h

A I D S . J I n f e c t D i s 1 9 8 8 ; 1 5 7 : 8 2 7 - 3 1 .

7 S e w a n k a m b o N , G o o d g a m e R , C a r s w e l l W J , e ta l . E n t e r o p a t h i c AI DS i n U g a n d a : a

m i c r o b i o l o g i c a l a n d h i s t o p a t h o l o g i c a l s t u d y .AIDS 1 9 8 7 ; 1 : 9 - 1 3 .

8 N i c h o l l s G . M i c r o s p o r i d i o s i s i n A I D S . C o m -

m u n i c a b l e D i s e a s e s R e p o r t C D R 8 6 / 2 1 . L o n -

d o n : P H L S , 1 9 8 6 : 3 - 4 .9 B r a n d b o r g L L , G o l d b e r g S B , B r e i d e n b a c h W C.

H u m a n c o c c i d i o s i s - a p o s s i b l e c a u s e o f

m a l a b s o r p t i o n . T h e l i f e c y c l e i n s m a l l - b o w e l

b i o p s i e s a s a d i a g n o s t i c f e a t u r e . N E n g l J Me d1 9 7 0 ; 2 8 3 : 1 3 0 6 - 1 3 .

V i s u a l a i d f o r q u i c k a s s e s s m e n t o f c o r o n a r ya r t e r y s t e n o s i s a t n e c r o p s y

A c h a r t w a s d e s i g n e d t o g i v e a q u i c k v i s u a la s s e s s m e n t o f t h e d e g r e e o f c o r o n a r y a r t e r ys t e n o s i s a t n e c r o p s y . I t w a s i n t e n d e d f o rr o u t i n e c a s e s i n w h i c h c o r o n a r y a r t e r ya n g i o g r a p h y a n d f o r m a l m o r p h o m e t r i c

a n a l y s i s o f p r o c e s s e d s e g m e n t s o f v e s s e l w e r e

n o t t o b e u n d e r t a k e n . T h e m e t h o d d e s c r i b e dh e r e i s n o t t h e m o s t a c c u r a t e , b u t i t i s

p r e f e r a b l e t o s i m p l e s u b j e c t i v e c a t e g o r i s a -t i o n i n t o s i m p l e , m o d e r a t e , a n d s e v e r e

d e g r e e s o f a r t e r y s t e n o s i s .I s c h a e m i c h e a r t d i s e a s e i s o n e o f t h el e a d i n g c a u s e s o f d e a t h i n G r e a t B r i t a i n .Q u a n t i f i c a t i o n o f c o r o n a r y a r t e r y s t e n o s i s a sp a r t o f t h e n e c r o p s y i s n e c e s s a r y t o e s t i m a t et h e f u n c t i o n a l i m p o r t a n c e o f a n y a t h e r o -s c l e r o t i c d i s e a s e p r e s e n t . T h e c r o s s s e c t i o n a la r e a o f t h e v e s s e l l u m e n , c o m p a r e d w i t h t h a tc o n t a i n e d w i t h i n t h e e l a s t i c l a m i n a , e x p r e s -s e d a s a p e r c e n t a g e , i s a w i d e l y u s e d m e t h o d

o f e s t i m a t i n g t h e d e g r e e o f l u m i n a l a r e an a r r o w i n g w i t h i n a v e s s e l . ' A n a r r o w i n g b y

75% r e d u c e s c o r o n a r y b l o o d f l o w a t t i m e s o f

s t r e s s a n d e x e r t i o n ; n a r r o w i n g b y 90%m e a n s t h a t c o r o n a r y b l o o d f l o w i s s e v e r e l yr e d u c e d a t r e s t . 2 E v e n i n t h e a b s e n c e o fc o r o n a r y t h r o m b o s i s , c a r d i a c d e a t h mayo c c u r i f t h e r e i s o n e s e g m e n t o f l u m i n a l a r e aw h i c h h a s n a r r o w e d b y m o r e t h a n 8 5 % ,

a l t h o u g h m o s t o f t h e s e p a t i e n t s h a v e m u l t i -p l e f o c i o f s t e n o s i s . 3

C o r o n a r y a n g i o g r a p h y c a n b e u s e d r o u t i n -e l y a t n e c r o p s y 4 t o s h o w t h e l u m i n a l d i a m e t e r

a n d i t c a n a l s o s h o w a l l b r a n c h e s o f t h ec o r o n a r y v e s s e l s . F o r m a l m o r p h o m e t r y w i t h

t h e u s e o f e i t h e r p o i n t c o u n t i n g , 5 o r p l a n -

i m e t r y o f m u l t i p l e c r o s s s e c t i o n s o f v e s s e l st h a t h a v e b e e n f i x e d , d e c a l c i f i e d , a n d s t a i n e df o r e l a s t i n , g i v e a g o o d e s t i m a t i o n o f l u m i n a l

a r e a n a r r o w i n g . V e s s e l s t h a t h a v e b e e n p e r -f u s e d a n d f i x e d a t p h y s i o l o g i c a l p r e s s u r e sb e f o r e d i s s e c t i o n g i v e t h e m o s t m e a n i n g f u l

r e s u l t s . 6 An a l t e r n a t i v e , b u t t h e m o s t r o u t i n -e l y u s e d m e t h o d , i s s i m p l e s u b j e c t i v e a s s e s s -

m e n t o f a s e r i a l l y s l i c e d v e s s e l a t t h e t i m e o fn e c r o p s y i n t o c a t e g o r i e s o f m i l d , m o d e r a t e ,

a n d s e v e r e s t e n o s i s .T h e f i r s t t w o m e t h o d s a r e o b v i o u s l y m o r e

a c c u r a t e , a n d t h e m e r i t s a n d d e m e r i t s o f e a c h

h a v e b e e n d i s c u s s e d e l s e w h e r e . ' T h e a i m o f

o u r m e t h o d i s n o t t o r e p l a c e o r c o m p e t e w i t h

t h e s e m e t h o d s b u t t o m a k e s i m p l e ,i m m e d i a t e , s u b j e c t i v e a s s e s s m e n t m o r e

a c c u r a t e a n d s c i e n t i f i c .E x a m i n a t i o n o f c o r o n a r y a r t e r i e s b y s e r i a l

t r a n s v e r s e s l i c i n g a t 3 mm i n t e r v a l s i s w i d e l yp r a c t i s e d . M o s t p a t h o l o g i s t s o p e n t h e h e a r tf i r s t a n d t h e n s l i c e t h e v e s s e l s w h i l e g r i p p i n g

t h e a r t e r y , w i t h f o r e f i n g e r o u t s i d et h e h e a r t

a n d t h u m b i n s i d e . A s h a r p b r a i n k n i f e a l l o w sa " c l e a n s l i c e " t o b e m a d e e v e n w i t h

m o d e r a t e v e s s e l c a l c i f i c a t i o n .O n c e t h e v e s s e l s h a v e b e e n s e r i a l l y s l i c e d ,

t h e c u t s u r f a c e i s c o m p a r e d w i t h t h e c h a r t( f i g u r e ) . T h e v e s s e l s h a v e b e e n d r a w n w i t h

c o n c e n t r i c r o u n d , e c c e n t r i c r o u n d , a n d s l i t

s h a p e d l u m i n a r e p r e s e n t i n g t h e t h r e e m a i n

p a t t e r n s s e e n a t n e c r o p s y . T h e d i a g r a m sh a v e b e e n p h o t o r e d u c e d f o l l o w i n g t h e c a l -c u l a t i o n o f t h e a r e a s o n g r a p h p a p e r .A r t e r i e s t h a t a r e d i s t e n d e d a t p h y s i o l o g i c a lp r e s s u r e a r e a l m o s t c i r c u l a r i n s h a p e . ' T h e

(@) -90%-0 -90%- (

( i # I ) - 3 0 % - K) - 6 0 - ((j)

-70%- -70%-

-60%- - 6 0 % -

-40%- -40%-

______ -20% -( -20%-

F i g u r e S c h e m a t i c d i a g r a m r e p r e s e n t i n g a

p e r c e n t a g e r e d u c t i o n i n c r o s s s e c t i o n a l a r e a

o f c o r o n a r y a r t e r y v e s s e l s . T h e o u t e r c i r c l er e p r e s e n t s v e s s e l e x t e r i o r , i n n e r c i r c l er e p r e s e n t s t h e e l a s t i c l a m i n a , a n d b l a c k a r e a

r e p r e s e n t s t h e l u m e n .

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8 8 8 L e t t e r s t o t h e E d i t o r

s l i t s h a p e s o f s o m e l u m i n a a r e a r t e f a c t u a ld u e t o c o l l a p s e : t h e d e g r e e o f s t e n o s i s a tn e c r o p s y i s t h e r e f o r e g r e a t e r t h a n i n l i f e , a n d

t h e d e g r e e o f c o l l a p s e w i l l v a r y a c c o r d i n g t ot h e p r o p o r t i o n o f t h e w a l l w h i c h i s f r e e o f

d i s e a s e ; t h i s c h a r t c a n o n l y b e u s e d a s a r o u g h

g u i d e i n t h e s e c a s e s . P r e s s u r e f i x a t i o n w o u l d

b e r e q u i r e d t o c i r c u m v e n t t h i s p r o b l e m .

R e q u e s t s f o r a c h a r t o n A4 s i z e p a p e r c a n

b e m a d e t o Dr C h a m p .CS CHAMP,*

S B COGHILL

D e p a r t m e n t s o f P a t h o l o g y ,* K e t t e r i n g a n d D i s t r i c t G e n e r a l H o s p i t a l ,

R o t h w e l l R o a d ,

K e t t e r i n g ,N o r t h a m p t o n s h i r e N N16 8UZ

N o r t h a m p t o n G e n e r a l H o s p i t a l ,C l i f t o n v i l l e ,

N o r t h a m p t o n N N I 5BD

R e f e r e n c e s

I D a v i e s M J . T h e p a t h o l o g y o f i s c h a e m i c h e a r td i s e a s e . I n : A n t o n y P P , M a c S w e e n RNM,e d s . R e c e n t a d v a n c e s i n h i s t o p a t h o l o g y 1 3 .E d i n b u r g h : C h u r c h i l l L i v i n g s t o n e , 1 9 8 7 : 1 8 5 -2 0 2 .

2 M a x i m i l i a n B u j a L , W i l l e r s o n J T . T h e r o l e o f

c o r o n a r y a r t e r y l e s i o n s i n i s c h a e m i c h e a r td i s e a s e . Hu m P a t h o l 1 9 8 7 ; 5 : 4 5 1 - 6 0 .

3 D a v i e s M J , P o p p l e A . S u d d e n u n e x p e c t e dd e a t h , a p r a c t i c a l a p p r o a c h t o t h e f o r e n s i cp r o b l e m . H i s t o p a t h o l o g y 1 9 7 9 ; 3 : 2 5 5 - 7 7 .

4 C o g h i l l S B , N i c o l l S M , McKimmie A , H o u s t o n

I , M a t t h e w BM. R e v i t a l i s i n g p o s t m o r t e m

c o r o n a r y a n g i o g r a p h y : J C l i n P a t h o l1 9 8 3 ; 3 6 : 1 4 0 6 - 9 .

5 M a n w a r r i n g L , O ' C o n n e l l D L , B h a g w a n d e e nB S , Z a r d a w i I M , D o b s o n A J . M o r p h o m e t r i c

a n a l y s i s o f c o r o n a r y a r t e r y s t e n o s i s ; a n

a c c u r a c y a n d r e l i a b i l i t y s t u d y . J P a t h o l

1 9 8 8 ; 1 5 6 : 1 1 1 - 1 7 .6 T h o m a s A C , D a v i e s M J . P o s t - m o r t e m i n v e s -

t i g a t i o n a n d q u a n t i f i c a t i o n o f c o r o n a r y a r t e r yd i s e a s e . H i s t o p a t h o l o g y 1 9 8 5 ; 9 : 9 5 9 - 7 6 .

7 A r n e t t E N , I s n e r J M , R e d w o o d D R , e t a l .

C o r o n a r y a r t e r y n a r r o w i n g i n c o r o n a r y h e a r td i s e a s e : C o m p a r i s o n o f c i n e a n g i o g r a p h i c a n d

n e c r o p s y f i n d i n g s . A n n I n t e r n Med1 9 7 9 ; 9 1 : 3 5 0 - 6 .

E l i m i n a t i o n o f p a r a l l a x e r r o r i n h a e m a t o c r i t

r e a d i n g s u s i n g r e f l e x i o n b a e m a t o c r i t o m e t r yT h e p r e c i s i o n o f h a e m a t o c r i t r e a d i n g s c a n b e

i m p r o v e d b y u s i n g a m a g n i f y i n g g l a s s . 'U n f o r t u n a t e l y , t h e p r e c i s i o n r e s u l t i n g f r o m

m a g n i f i c a t i o n m a y b e u n d e r m i n e d b y t h ep a r a l l a x e r r o r . T h e c o n s e q u e n c e o f t h ep a r a l l a x e r r o r i s e a s i l y r e c o g n i s e d i f t h e e y eo r t h e l e n s i s m o v e d a l o n g t h e a x i s o f t h em i c r o h e m a t o c r i t t u b e ; t h e r e a d i n g s o f t h ee r y t h r o c y t e s , b u f f y l a y e r a n d p l a s m a c o n c e n -

t r a t i o n w i l l b e c o m p l e t e l y c h a n g e d . T h e

r e s p e c t i v e e r r o r o f t h e h a e m a t o c r i t i n c r e a s e sw i t h d e c r e a s i n g h a e m a t o c r i t a n d c y t o c r i t .

To a v o i d t h e p a r a l l a x e r r o r , a r e f l e x i o nh a e m a t o c r i t o m e t e r w a s d e v e l o p e d a c c o r d i n gt o t h e f o l l o w i n g p r i n c i p l e . A t r a n s p a r e n ts c a l e w a s m o u n t e d o n a s u r f a c e m i r r o r a n d

c o v e r e d b y a g l a s s l a y e r . T h i s r e f l e c t i n g u n i tw a s p r o v i d e d w i t h a s c r e w d r i v e n f i x a t i o nd e v i c e f o r t h e m i c r o c h e m a t o c r i t t u b e s o t h a tt h e b o t t o m o f t h e r e d c e l l c o l u m n c o u l d b e

g e n t l y a d j u s t e d t o t h e z e r o l i n e o f t h e s c a l e( f i g u r e ) . When o b s e r v e d b y a m a g n i f y i n g

g l a s s , t h e r e a l a n d r e f l e c t e d c e l l a n d p l a s m a

l a y e r s c a n b e a d j u s t e d t o a common l e v e l ,i n d i c a t i n g t h e r e a d i n g p o s i t i o n o f t h e e y e a n dt h e l e n s , t h u s a v o i d i n g t h e p a r a l l a x e r r o r .C o e f f i c i e n t s o f v a r i a t i o n w e r e a s s e s s e d w i t h

w h o l e b l o o d a n t i c o a g u l a t e d w i t h e d e t i c a c i do r l e u c o c y t e p o o r e r y t h r o c y t e s u s p e n s i o n s i na n t i c o a g u l a t e d p l a s m a o r i n b u f f e r .

T e n m i c r o h a e m a t o c r i t c a p i l l a r i e s o f

7 5 mm i n l e n g t h w e r e f i l l e d t o a 6 0 mm m a r k

a n d s e a l e d . B e t w e e n e a c h f i l l i n g p r o c e d u r et h e b l o o d t u b e w a s c l o s e d a n d m i x e d b y

h a n d . T h e 1 0 c a p i l l a r i e s w e r e c e n t r i f u g e dt o g e t h e r f o r f i v e m i n u t e s ( 1 2 5 0 0 x g a t t h eb o t t o m o f t h e t u b e s ) . A f t e r c e n t r i f u g a t i o nt h e y w e r e k e p t i n v e r t i c a l p o s i t i o n u n t i l t h er e a d i n g . I n e x p e r i m e n t s 8 - 1 7 r e p e a t e d r e a d -i n g s o f s i n g l e c a p i l l a r i e s w e r e p e r f o r m e d .I n e x p e r i m e n t s 1 - 1 6 h a e m a t o c r i t m e a s u r e -

m e n t s w e r e p e r f o r m e d w i t h t h e i n s t r u m e n tp l a c e d h o r i z o n t a l l y ; e x p e r i m e n t 1 7 w a s d o n ew i t h t h e i n s t r u m e n t i n v e r t i c a l p o s i t i o n . A1 0 x e y e p i e c e o f a m i c r o s c o p e w a s u s e d f o ra l l t h e r e a d i n g s b e c a u s e t h i s i s a v a i l a b l e i nm o s t h a e m a t o l o g i c a l l a b o r a t o r i e s . T h e t a b l e

s h o w s t h e c o e f f i c i e n t s o f v a r i a t i o n o f th a e m a t o c r i t o f w h o l e b l o o d a n d o f e r y tr o c y t e s u s p e n s i o n s i n p l a s m a o r b u f f e r . Ah i g h c o n c e n t r a t i o n s o f h a e m a t o c r i t a l l tc o e f f i c i e n t s o f v a r i a t i o n w e r e a r o u n d

b e l o w 0 - 5 %. F o r t h o s e a r o u n d 0 0 7 , tc o e f f i c i e n t s o f v a r i a t i o n a v e r a g e d 2 5 % .e x p e r i m e n t 1 t h e b u f f y l a y e r w a s r e a da d d i t i o n t o t h e p l a s m a a n d r e d c e l l l a y e r s a n

t h e c y t o c r i t ( s u m o f t h e r e d p l u s w h i t e c ec o l u m n s d i v i d e d b y t h e l e n g t h o f t h e w h o l

c o l u m n ) w a s a l s o c a l c u l a t e d a n d w a

0 - 4 7 8 ± 0 0 0 1 a n d t h e c o e f f i c i e n t o f v a r it i o n 0 - 2 1 % . I n e x p e r i m e n t s 8 - 1 7 , 1 0 r e p e a tr e a d i n g s o f s i n g l e c a p i l l a r i e s w e r e p e r f o r m eI n e x p e r i m e n t s 8 - 1 3 c o e f f i c i e n t s o f v a r i a t iw e r e o b t a i n e d s i m i l a r t o t h o s e i n e x p e r i m e n

1 - 7 . T h i s w a s d u e t o t h e f a c t t h a t m a n i p u l

t i o n b e t w e e n r e a d i n g s l e d t o s o m e c r o w d i

o f t h e u p p e r m o s t e r y t h r o c y t e l a y e r a f t e r f ot o s i x r e a d i n g s ( t a b l e ) . I n e x p e r i m e n t s w h e r

m a n i p u l a t i o n w a s r e d u c e d t o t h e n e c e s s a

m i n i m u m , t h e c o e f f i c i e n t o f v a r i a t i o n w az e r o f o r w h o l e b l o o d a n d e r y t h r o c y t e s s u se n d e d i n b u f f e r a t a l o w h a e m a t o c r( e x p e r i m e n n t s 1 4 , 1 5 ) . A t a h i g h e r c o n c e n t rt i o n o f e r y t h r o c y t e s i n b u f f e r , h o w e v e r , s o m

c r o w d i n g a l s o o c c u r r e d ( e x p e r i m e n t 1 6 ) b

c o u l d a l m o s t b e s u p p r e s s e d w h e n t h e i nt r u m e n t w a s i n v e r t i c a l p o s i t i o n ( e x p e r i m e1 7 ) .

T h e h i g h p r e c i s i o n o f t h e r e a d i no b t a i n e d w i t h t h i s i n s t r u m e n t c o m p a r ew i t h t h o s e o b t a i n e d w i t h a n o n - r e f l e c t is c a l e w a s d u e t o ( 1 ) t h e e l i m i n a t i o n o f tp a r a l l a x e r r o r a n d ( 2 ) t h e m e c h a n i c a l a d j u s

m e n t o f c a p i l l a r i e s t o t h e i n s t r u m e n t s c a l

- ' .~ ..B....

. . J c _ . . . . . . . . . . . . . . . . . . . . . . . . . .F i g u r e R e f l e x i o n h a e m a t o c r i t o m e t e r w i t h h a e m a t o c r i t c a p i l l a r y a n d e y e p i e c e i n p o s i t i o n a

s c r e w ( a ) f o r d r i v i n g t h e c a p i l l a r y h o l d e r ( b ) a n d l i g h t s o u r c e ( c ) .

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C S Champ and S B Coghill

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