40th SEMI-ANNUAL SEMINAR (Fall 1972)
CALIFORNIA ASSOCIATION OF CRIMINALISTS
October 19-21, 1972
SACRAMENTO, CALIFORNIA

MICROSCOPY SEMINAR - Equipment, Operation and Maintenance
James R. Carson, Optical Specialist, Van Waters and Rogers; Ivan Z. Burba, Factory Representative, Carl Zeiss, Inc.

Mr. Carson: A general review of mechanical and optical parts of the two basic types of microscope. Mr. Carson emphasized the need to choose compatible components to achieve the highest resolution. The consequences a microscopist must face when he attempts to increase or decrease resolution, depth of field, contrast, or the intensity of illumination were discussed.
Mr. Burba: A brief review of phase contrast, polarized light, and fluorescence microscopy.


THE NEW LOOK IN MICROSCOPY FOR FORENSIC SCIENCES
John L. Yee, General Electric Company, Vallicitos Nuclear Center

The introduction to the paper covers briefly the history, significant developments and the era of ultramicroanalysis. A discussion concerning the use of morphological analysis with the visible light microscope, techniques of polarized light microscopy, dispersion staining, refractive index determination and limits of sensitivity is presented with emphasis toward application in the criminalistics areas. The development of the clean-bench for use in optical microscopy is outlined and its applications in Forensic Science problems is presented. Suggestions for application are in the areas of analysis involving microscopic tests with physiological fluid, narcotics, crystal and color tests and areas of trace evidence examination where contamination is a problem. Techniques in handling small amounts of material such as glass, pollen, fibers. Insect parts, paint and other materials encountered in evidence is discussed with reference to recognized experts who have been responsible for this type of technique development. The paper goes on to describe the information available from each sample when analyzed by the instruments with an inherent capability for small sample analysis such as the electron microprobe, transmission electron microscope, scanning electron microscope and ion microprobe mass analyzer. Information pertaining to elemental analysis, topography, quantitative aspects, sensitivity and non-destructive nature is given with reference to available information. A challenge is issued to the Criminalist to develop the use of the techniques and instruments in the examination of Forensic trace evidence.


A RAPID MICRO-METHOD FOR TYPING OF THE GENETIC VARIANTS OF THE ISOENZYMES OF PHOSPHOGLUCOMUTASE
Benjamin V. Grunbaum, Environmental Physiology Laboratory, University of California

A micromethod for typing human blood for the genetic variants of the isoenzymes of phosphoglucomutase was described. The procedure utilizes cellogel, which is a gel form of cellulose acetate, as the supporting medium for electrophoresis and an apparatus designed by Grunbaum et al. (Microchem. J. 7: 41-53, 1963). A buffer system consisting of Trisglycine at pH 8.4 permitted the use of high voltage (500 volts, for fast separation) and minimal current (about 2 m.amp. to avoid overheating). The hemolysate was applied close to the cathode in 10 applications of 0.25 microliters. After 1 hour electrophoresis, the membrane was placed face side down into a 1% noble agar reaction mixture and incubated in the dark at 37C. The isoenzymes became visible after 30 to 60 minutes. The 60 minute electrophoresis gave an adequate resolution and the hemoglobin band did not interfere. Also, the intensity of the minor isoenzymes was sufficient for type recognition. The electrophoretograms were recorded by color photography as soon as the pattern was complete. Delay in photography results in excessive intensification of the Individual components and reduction in contrast with the background agar gel. Work is underway to further improve the sensitivity of the method, through the use of regular cellulose acetate, which will require less sample applications.


A FAST AND SIMPLE CROSS-OVER ELECTROPHORESIS TECHNIQUE FOR IDENTIFICATION OF BIOLOGICAL STAINS
Benjamin W. Grunbaum

At this time gel media are used for most immunochemical reactions, including also cross-over electrophoresis (COE). It was recently pointed out by the author (J. Forens. Sci. Soc. (1972) 12, 431) that gels, being optically clear, are useful in visually following the course of development of inmunoprecipitin bands to any desired degree of completion. However, a number of difficulties inherent in the use of gels were also listed. Additional disadvantages in the use of gels, especially for COE, were found to be:

  1. The relatively long time required to fill a series of pairs of wells with antigen and corresponding antisera. Since COE is not only a YES or HO type of analysis, but also one of comparison, it appeared that considerable diffusion in 360 directions occurred, thereby diluting the reactants in the first wells by the time the reagents were placed in the last wells. The Hyland plates for Instance have 24 pairs of wells and it requires an experienced analyst about 30 minutes to fill them.
  2. The sample requirement for both the antigen and antiserum is rather large. It normally requires 5 to 7 microliters to fill each well. In this fashion, the reactants, which are scarce and expensive, are readily used up.
  3. The filling of tiny wells within gels is frustrating because underfilling, overfilling, or nicking of the gels causes artifacts.
For the above reasons, an applicator was designed which applies 8 samples at a time onto a cellulose acetate supporting medium. Since cellulose acetate is a more sensitive medium than gel, the sample requirement can be reduced about 20 fold. Also, since very dilute buffers can be utilized and thus a high voltage applied across the membrane, COE precipitin bands are complete within minutes. Best results were obtained by applying the reactant samples in the form of narrow bands. The precipitin bands thus obtained were straight and intense. The 8-sample applicator was integrated into the electrophoretic apparatus in a manner that made it possible to place the reactants at any desired proximity to each other and also with respect to the cathode and anode. No reference marks were needed on the cellulose acetate membrane itself.

FABRIC FLAMMABILITY AND TESTING PROCEDURES
Gordon Damont, Textile Chemist, California Bureau of Home Furnishings

The hazard associated with the combustibility of textile products has been of serious concern to man ever since he learned to use fire for constructive purposes. The State of California has been a leader in developing test methods for measuring flammability of apparel and for flammability testing in general. Current methods include the 45, one second test, "Timed Burning Tablet," and the timed vertical burn using 3- and 12-second open flame. Cigarettes are used as ignition when mattresses are being tested for flammability, but are difficult to control due to difference in ember temperatures.


AN ANALYTICAL APPROACH TO ULTRA-SMALL CRIME SCENE EVIDENCE
Victor Reeve, California State Bureau of Investigative Services; Arvid Lads, University of British Columbia

Electron microprobe analysis being a nondestructive, quantitative method of instrumental analysis is most suitably applied to problems encountered in forensic sciences. The added benefit of the examination of ultra-small specimens by this method enables the criminalist to extend analysis to "scene of crime" materials that previously could not have been considered on a quantitative basis.


FURTHER BLOODSTAIN STUDIES WITH THE AUTOANALYZER
Robert R, Ogle, Contra Costa County Sheriff's Laboratory

A history of the use of the autoanalyzer by the Contra Costa County laboratory for the determination of Rh factors was presented. Additional work in determining the A subgroups with the autoanalyzer was described, including a discussion of some problems encountered with older stains and with fresh stains.


THE FIELD DIFFERENTIATION OF HUMAN AND ANIMAL SKELETAL REMAINS
Robert R. Ogle, Contra Costa County

A brief discussion of the differences in certain skeletons was presented to assist criminalists in the field determination of skeletal remains of non-human origins.


A RAPID SIMPLIFIED METHOD FOR BARBITURATE ANALYSIS BY T.L.C.
Donald W. Jones, M.D.; De Witt Hunter, M.D.; George Shirman, M.T., Jones Medical Laboratory

When barbiturate levels are needed for emergency diagnostic purposes, the clinical laboratory technician does not have time to prepare fresh standards to run as knowns on a T.L.C. system. The authors' laboratory has developed a method where the standards are stored on discs for extended periods of time. The authors demonstrated the application of the technique to forensic problems by exhibiting an extraction flask which traps the questioned material on a similar disc. The discs fit neatly into a prepared T.L. sheet. The authors have prepared discs of known barbiturates, stimulants, tranquilizers, narcotics and general toxic mixtures.


CRIME LAB, CLARK COUNTY SHERIFF'S DEPARTMENT, LAS VEGAS, NEVADA
Wes Summerfield

With the exception of fingerprint and ballistic-type types of examinations, the dark County Sheriff's Department's Crime Lab services, equipment, routine and semi-routine methods of analysis and some interesting statistics concerning work load and expected levels of performance, both from a standpoint of time and thoroughness, are presented.


A SPECIFIC MICROCRYSTALLINE TEST FOR L.S.D.
Paul J. Cashman, Contra Costa Sheriff's Laboratory

A preliminary report was presented demonstrating the crystals developed after dissolving pure L.S.D. in 10% HC1 followed by the addition of a drop of 15% Ethylenediamine. Clean-up by solvent-solvent extraction or TCL was suggested for mixtures or dirty samples. Resulting pure material should approximate 50 micrograms. Crystals are examined by polarizing microscope. To date, the L.S.D. crystal can be distinguished from the other ergot alkaloids studied.


IDENTIFICATION OF CUT ENDS OF MULTI-STRAND WIRES
Robert R. Ogle and Gerald T. Mitosinka, Contra Costa Sheriff's Laboratory

A case presented to the author as described in which the issue was to determine if several wire segments had been previously joined. The distortion on the cut ends precluded identification by a classical physical match. The identification was effected by preparing a silicone rubber (Jelcon) cast of the insulation interior of the multi-strand wires. The casts were compared under a bullet comparison microscope. The patterns produced in the plastic insulation by the impressions of the wires were found to correspond exactly between pairs of segments, allowing an identification to be effected.


LOCATION OF SEMINAL STAINS BY THEIR PHOSPHORESCENCE
Peter Jones, G. Stupran, A.R. Galloway, & S. Siegel, Aerospace Corporation

Ultraviolet lamps have been used in the past for locating seminal stains by their blue fluorescence. Unfortunately, the widespread use of fluorescent additives in detergents and some clothing fibers has limited the usefulness of this technique. We have found, however, that seminal stains are readily located by cooling the sample and viewing its phosphorescence (defined below) instead of its fluorescence. This method of viewing the semen is also useful for determining the order of deposition of overlapping blood and seminal stains. The order of deposition of blood and seminal stains, at least for the type of fabric and pattern of deposition used to prepare the specimens, can be determined by examination of the blood and semen flow patterns if the interval between deposition of the stains is not too short. Both examination under visible light and observation of low temperature phosphorescence are useful complimentary techniques. For semen spreading into dried blood, order of deposition can certainly be resolved if the interval of deposition is at least 45 minutes, and probably for an interval of at least 15 minutes. For blood spreading into dried semen, order of deposition may be ascertained if the interval between depositions was at least 15 minutes.


COMPARATIVE MERITS OF THE ACID PHOSPHATASE TEST FOR SEMINAL STAINS
Robert Ogle, Charles Smith, Contra Costa Sheriff's Laboratory; John Thornton University of California, Berkeley

A brief discussion of the relative merits of the acid phosphatase test for semen stains was presented. The "specificity" of the test was discussed, including the use of the term as it applies to the determination of the absence as well as the presence of semen. A new substrate for the acid phosphatase test was described which possesses greatly enhanced specificity for the prostetic enzyme. Data were presented (1) to indicate the types and frequencies of tests for semen utilized by California laboratories, (2) to indicate the results obtained in 100 semen examinations in the authors' laboratory, and (3) to, indicate the relative specificity of the acid phosphatase test as it is currently employed, and with the new substrate (sodium thymolphthalein monophosphate).


A BRIEF LOOK AT DTA, OXYLUMINESCENCE AND THERMOLUMINESCENCE
J.D. DeHaan & J.M. Rynearson, Alameda County Sheriff's Laboratory

A study of thermal analysis reveals a long history of various analytical techniques being used to gain a maximum of information from small quantities of many different materials. The combination of several productive thermal methods, Differential Thermal Analysis (DTA), Thermolumin-escence (TOL), and oxyluminescence (OxyL), was accomplished here in a single, relatively simple unit. The experimental apparatus consists of the traditional elements: sample block with axial heater, sample and reference wells, and thermocouples, all contained in an aluminum chamber to provide a controlled environment. For luminescence studies, open sample pans are used, and a sensitive photomultiplier tube monitors sample behavior. The innovative use of solid-state electronic circuitry permits great flexibility in design application. DTA is useful for revealing crystal states, hydration states, and thermal history of small (ca. 20 mg) samples of such materials as plaster, explosives, and inorganic salts. It reveals such fundamental properties as melting and boiling points and thermal decomposition characteristics and is very sensitive to the presence of impurities. In many cases, materials are not decomposed by DTA, and subsequent IR, UV or spectro-graphic analyses can be performed. Thermal luminescence studies permit characterization of a material by an examination of its light-emitting behavior at various temperatures. Thermoluminescence can reveal a sample's purity and thermal history as well as characterize it, and typically uses 5-20 mg of solid, non-conducting materials as glass and inorganic salts. Oxyluminescence permits characterization of small (l mg) samples of polymer fibers and films such as Nylon, Teflon, poly-vinyl chloride and cellulose acetate as they are oxidized at elevated temperatures, and reveals the presence and effects of anti-oxidants and catalysts. Although thermal methods are generally unsuitable for specific identifi-cations, they are shown to be extremely useful for characterization and comparison of parameters between two or more samples. In many cases, the effects which are so vividly displayed in thermograms or glow curves are not revealed by any other instrumental or chemical techniques.


ABSTRACT OF REPORT ON TRANSLATION OF FOREIGN CRIMINALISTIC ARTICLES
Herman J. Meuron

Until recently there has been no systematic search or translation of foreign articles in the field of Criminalistics done by the National Technical Information Service. Now, LEAA, through Joseph Petersen, Criminalistics Specialist, is exploring the possibilities of funding such a translation program through the National Science Foundation. Yet to be finalized are the exact articles to be translated and the distribution.


ABSTRACT OF REPORT ON THE FORENSIC SCIENCE SYMPOSIUM OF THE ASSOCIATION OF OFFICIAL ANALYTICAL CHEMISTS HELD IN WASHINGTON, D.C., OCTOBER 12, 1972
Herman J. Meuron

An introductory paper, a General Referee's Report, and. a scientific paper were covered dealing with forensic aspects of and applications of voice prints, neutron activation analysis, atomic absorption, electron microscopy, chemical ionization mass spectrometry, laser beam mass spectrometry, microscopic techniques for glass analysis, vapor trace analysis of explosive residues, and methods for drug and drug adulterants.