TB ALMOST WHILE YOU WAITHITHERTO cultivation of tubercle bacilli has been too
low a process to be of much use in diagnosis, but D. M.Pryce has described a micro -culture method which yieldswell-grown cultures within a week. Indeed, obvious signsof growth are visible with the TIf in. objective in 24 hours,colonies may be just perceptible with the in. objectivein 48 hours and in 3 days they are usually quite distinct.The method consists in drying a film of sputum on a glasssurface, treating with 15% sulphuric acid, washing withwater and incubating with blood hsemolysed with dis-tilled water or 1% saponin. After 7 days incubationthe preparation is washed and stained with Ziehl-Neelsen and the colonies examined with the low power.The most typical colonies are long and stringy, laterbecoming twisted and coiled, but compact and bushyforms occur. Mycelial threads are found, and there issome suggestion that growth does not take place through-out the length of the thread but is restricted to certainpoints. Two methods of applying this dried-film culturetechnique are described. In one the sputum is spread onthe floor of a petri dish. In the other one or more cir-cular patches of sputum are spread on a glass slide, dried,surrounded with a ring of bakelite, hard fibre or glassimmersed in melted soft paraffin, and incubated in amoist chamber or with a coverslip to cover the ring.The method was tried out on sputa from 48 cases fromthe male wards of Harefield Sanatorium. In 28 growthwas luxuriant and immediately obvious, and in 6 coloniesgrew which were sparsely distributed. The remaining14 yielded no growth. The technique can also be appliedto pus and caseous material.
PLASMA IRONADVANCING biochemistry now allows the iron in blood
plasma to be measured. When every precautionagainst haemolysis is taken there still remains a minutebut fairly constant amount of iron measured in micro-grammes per c.cm. Vahlquist 2 gives the normal limitsas 80-190 pg. per 100 c.cm. in men and 60-190 ug.in women with average figures of 122 and 104. Indisease these levels differ ; but if we are to apply thisnewly acquired technique we must understand what hasrecently been learnt about the absorption and transportof iron. Hahn, Whipple and their co-workers 3 used"
marked " radio-active iron to investigate this problem.They found, as Widdowson and McCance 4 had done,that iron absorption is proportional to iron needs andthat once it is absorbed there is little excretion of iron.When iron salts are fed to dogs rendered ansemic bybleeding the radio-iron in the plasma rises rapidly,reaching a peak within 4-8 hours and then rapidlyfalling to resting level in 6-12 hours after feeding.Radio-iron appears in red blood-cells in traces as early as4 hours after feeding and after 24 hours is present inconsiderable amounts. Only about 10% of the absorbediron can be accounted for as haemoglobin iron, the restpresumably goes to iron stores. The plasma ironthus represents iron passing between the alimentarytract, the red-cell forming tissues and the iron stores,and we might expect that the response to atest doseof iron would give some indication of the presence orabsence of iron-deficiency. Vahlquist gives a series of" iron-tolerance curves " in various conditions. In anormal person the test dose causes a rise of serum ironto about 300 /kg. per 100 c.cm. in 4-6 hours, with returnto normal level in 12 hours when fasting. The serum1. J. Path. Bact. 1941, 53, 327.2. Vahlquist, B. C. Acta pdiatr., Stockh. suppl. 5, 1941.3. Hahn, P. F., Whipple, G. D. et al. J. exper. Med. 1939, 69, 793;
70, 443.4. Widdowson, E. M. and McCance, R. A. Biochem. J. 1937, 31,
iron is below normal in anamia after haemorrhage, simplehypochromic anaemia of adults and children, and anaemiaaccompanying infection. In the first two a test dosecauses a large rise in serum iron and after 12 hours thefasting level has not been regained ; in the infectionanaemia there is little response. In pernicious anaemiathe fasting level may be as high as 360 flg. per 100 c.cm.and a test dose of iron causes little further increase ;after liver treatment the resting serum iron falls tonormal or below, and some iron deficiency is not un-common during the remission period. The fasting levelis also high in hsemolytic anaemias. Waldenstrom 5thinks that in clinical work serum-iron determinationwill be most useful in detecting iron deficiency. He hasdescribed patients with epithelial symptoms, such assore mouth and dysphagia, but no anaemia, who havebeen shown to be deficient by the iron-tolerance test andwho have responded satisfactorily to large doses of iron-so-called" latent sideropenia." The test is also useful indetecting iron-deficiency in patients with a normo-chromic anemia, in the remission phase of perniciousansemia, and after gastric operations. There is onemore finding that may prove useful : in acute hepatitisthe serum iron may be raised as high as 370 p,g. per100 c.cm., whereas in obstructive jaundice it isunaffected. Serum-iron determination then maytake its place in the long list of biochemical tests fordifferentiating toxic and obstructive jaundice..
This sort of technique has also been used to estimatethe value of the various iron preparations now used
therapeutically. A test dose containing a standardquantity of iron is given and the change in serum ironfollowed. Moore and others,6 and Vahlquist, workingon human subjects, found that ferrous salts were bestabsorbed, the rise in serum iron being unaffected by theabsence of HCI in the stomach. Ferric salts, togetherwith reducing agents like ascorbic acid or sodium formal-dehyde sulphoxylate, gave figures equal to ferrous iron ;ferric salts alone and other iron preparations wereinferior. They noted that food influenced the absorptionof iron, a subject discussed by Tompsett 7 who thinksthat iron is only absorbed in the ferrous state.Hahn, Whipple and their colleagues,8 in their experi-
ments with anaemic dogs on an iron-free but otherwisenormal diet, got different results. They found the largerthe doge of iron the smaller the percentage absorbed, sothat it was best to give small divided doses. Whenconditions were optimal absorption was just as rapidwhether the iron was given as a ferric or ferrous salt oreven in an organic combination. The rate of utilisationof the absorbed iron was increased by previous ironfeeding and by a diet of raw lean beef. By far the bestresults were obtained by intravenous injection of colloidaliron. How far these results on normal, temporarilyanaemic dogs are applicable to abnormal anaemic men it isdifficult to say. The use of intravenous iron suggeststhat a really efficient preparation to replace the unsatis-factory injectio ferri (BP) is needed. Enough has beensaid to show the usefulness of plasma or serum irondeterminations. The methods available are, however,not simple, and for accuracy demand scrupulous cleanli-ness, iron-free reagents, and apparatus outside the usualrange of laboratory equipment. The most favouredmethods are based on the technique of Heilmeyer andPlbtner.9 A protein-free filtrate is prepared and thecolour reaction between the iron in this filtrate and asubstance like o-phenanthroline is used for the estimation.A photo-electric colorimeter or similar electrical instru-ment is necessary for accuracy. With these difficultiesserum iron estimations are not likely to be popular, but
5. Waldenstrm, J. Nord. Med. 1941, 11, 2341.6. Moore, C. V. et al. J. clin. Invest. 1939, 18, 553.7. Tompsett, S. L. See Lancet, 1940, ii, 365.8. J. exper. Med. 1940, 71, 731.9. Heilmeyer, L. and Pltner, K. Das Serumeisen, Jena, 1937.