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Rubber dam clamps are known to break during clinical use in endodontics. This in-vitro study examined some of the variables which may contribute to the fracture. Stainless steel rubber dam clamps were subjected to various cleaning and autoclaving regimes and exposure to various solutions of sodium hypochlorite (NaOCl). Each clamp was examined after four cycles of cleaning and exposure to NaOCl. During environmental exposure to NaOCl, the clamp was stressed over a perspex rod to simulate placement onto the crown of a tooth. Clamps were examined after each test cycle visually and microscopically, or immediately after breakage. Results suggested that the fractures were because of a stress corrosion cracking phenomenon. There was evidence of intergranular and transgranular cracking of the metal. Corrosion spots were seen on the surface of the clamps and fracture occurred mainly through these spots. A number of recommendations to reduce breakage of clamps have been suggested.

The existence of 4 alleles of phosphoglucomutase (PGM1) in human red cell lysates has been demonstrated by isoelectric focusing confirming earlier work by Bark et al. and Kuhnl et al. A survey of 101 red cell lysates and the inheritance of these alleles in 24 families are now described.

A preliminary study has been made of the use of immobilised pH gradients (Immobiline gels) for the separation of the six commonly occurring erythrocyte acid phosphatase (ACPl) phenotypes in blood lysates and stains. These pH gradients are immobilised in the matrix of the polyacrylamide gel and possess different properties from conventional isoelectric focusing (IEF) gels. A comparison was made between the profile of the ACP 1 isoenzymes apparent on conventional IEF gels with that seen on the Immobiline gels. I IntroductionHopkinson et al. [ Although IEF has proved to be a more sensitive method oftyping ACPl activity than the starch gel technique, particularly in bloodstains, there are still problems in interpretation, paricularly where A activity is low and where one has to discriminate between the B and CB phenotypes. Recently immobilised pH gradients have been described [7,81 which are prepared with a set of commercially available acrylamide derivatives (Immobiline). Both basic and acidic Immobilines are mixed separately with a number of reagents normally used in the preparation of polyacrylamide gels in a gradient mixer to form a gel which varies in pH. The Immobilines finally used will depend upon the pIrange ofthe system to be investigated, which, for the common ACPl variants, is pH 5.77-7.58 [91. With these considerations in mind a study was set up to evaluate the Immobiline technique with particular regard to the problems associated with ACP 1 typing in bloodstains. This preliminary report follows the separation of six common phenotypes of ACPl from blood lysates and stains using the Immobiline gel system and discusses the advantages of this technique over conventional IEF. Materials and methods Preparation of red cell lysates and bloodstainsBlood was collected in heparinized tubes by finger pricks from known ACPl donors. After centrifuging, the serum was removed and the erythrocytes were washed twice with 0.85 % wlv saline between further centrifugations. The resultant pellet of erythrocytes were then lysed by freezing and thawing. Bloodstains were made from finger pricks on cotton cloth from donors with known and unknown ACP 1 phenotypes, and were air-dried before analysis. Immobiline gel preparationGeneral details of gel preparation are given in LKB Application Note 32 1 [71. The gels were 245 x 100 x 0.5 mm and, for sample application, they contained slots of the size 10 x 2 x 0.2 mm, made by applying Dymo Tape to the surface of the glass plate. A pH gradient of pH 5.5-7.6 was selected which would give the optimal separation ofthe commonly occurring ACPl phenotypes which have PI values in the range pH 5.77-7.58 191. Stock solutionsAcrylamide 29.1 % w/v containing bisacrylamide 0.9 % w/v; glycerol 87 % w/v; Immobilines 0.2 M solutions of pK 7.0, pK 6.2 and pK 3.6; ammonium persulphate 10 % w/v; NNN'N'-tetramethylethylenediamine (TEMED) 10 % v/v (Table 1).

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