1. ‫العظي‬ ‫هللا‬ ‫صدق‬‫م‬
‫االسراء‬ ‫سورة‬(85)

2. Role of laboratory in
Toxicological Studies
Prepared by
Reda M. Elsayed
29/4/2015

4. Types of Biological samples:
1- BODY FLUIDS:
•Blood (plasma, serum and whole
blood)
•Urine, semen, saliva, gastric
contents..etc.
2- Tissue samples:
liver, kidney, muscles….etc and
Hair.

8. Epindorf tubes
micropipettes

9. a) Biochemical and
toxicological study
Homogenized by Tissue
Homogenizer/ Grinder

12. b) Histopathology
exam.:
Organs Preserved
as it is in different
preservative
according to type
of tissue then
processed by the
specialist.

13. a- Biochemical Tests.
b- Toxicological Tests.
c- Histopathological Exam.

14. 1. Spectrophotometery:
Used for liver and kidney functions,
glucose, pseudocholine
estrase...etc
2. Immunoassay ( ELISA and RIA):
Hormones and some enzymes….etc.

15. 1. Spectrophotometery
2. Immunoassay ( ELISA and
RIA):
3. Chromatography:
 TLC
 HPLC, LC- MS
 GC, GC-MS

17. Bechman chemistry
apparatus.

18. •“Immuno” refers to an immune response that
causes the body to generate antibodies.
•“Assay” refers to a test.
Immunoassays are chemical tests used to
QUALITATIVE or SEMIQUANTITATIVE to
determine a specific substance, the analyte, in
a blood or body fluid sample, using an
immunological reaction.

19. RIA
In which, there is a competition between two antigens:
the labeled antigen reagent (Ag*) and
the unlabeled specimen (Ag) (or test sample analyte)
Both compete for a limited amount of antibody.

20. Tramadol PROCEDURE
The assay is based on the competition of Tramadol
labeled enzyme (G6PDH) and the free drug in the
urine sample for the fixed amount of antibody
binding sites.
In presence of the drug, The enzyme (G6PDH) is
free so activity is determined by the conversion of
NAD to NADH…. Colour change …positive test.
In the absence of the free drug in the sample, the
antibody binds the drug enzyme conjugate and
enzyme activity is inhibited.

21. Drug analyzer
Immunoassay

22. ELISA
An enzyme-linked immunosorbent
assay (ELISA) used to detect the
presence and/or amount of a target
protein of interest within an
experimental sample.
Detection of the target protein is
made by antibodies, which make the
ELISA an immunoassay.
Through a series of incubation and
washing steps.

24. Architect plus I 1000
Immuno- analyser

27. Liquid-liquid extraction is a useful
method to separate components
(compounds) of a mixture

28. For example:
Suppose that you have a mixture of sugar
in vegetable oil (it tastes sweet!) and you
want to separate the sugar from the oil.
You observe that the sugar particles
are too tiny to filter and you suspect that
the sugar is partially dissolved in the
vegetable oil.

29. How about add water to the
mixture
Will it separate the sugar from the
oil? Sugar is much more soluble in
water than in vegetable oil, and, as
you know, water is immiscible (=not
soluble) with oil. ‫هل‬‫يختلط‬‫الزيت‬
‫بالماء‬
The water phase is the
bottom layer and the oil phase
is the top layer, because
water is denser than oil.

30. By shaking the layers
(phases) well, you
increase the contact
area between the two
phases. The sugar will
move to the phase in
which it is most
soluble (the water
layer)

32. In this example:
water was the extraction solvent.
the original oil-sugar mixture was
the solution to be extracted.
sugar was the compound (analyte)
extracted from one phase to
another.

33. the concept of liquid-liquid extraction:
Liquid-liquid extraction is based on the
transfer of a solute substance from one
liquid phase into another liquid phase
according to the solubility.
You can use extraction to separate a
substance selectively from a mixture, or
to remove unwanted impurities from a
solution.

34. In the practical use, usually one phase
is a water or water-based (aqueous)
solution and the other an
(organic solvent) which is immiscible with
water.
The success of this method depends
upon the difference in solubility of a
compound in various solvents. And the
choose of a suitable extraction solvent.

35. 1. Thin layer chromatography
2. Liquid Chromatography
3. Gas Chromatography (GC)

36. Chromatographic separation process is
based on the difference in the surface
interaction of the analyte (sample) and
eluent molecules (mobile phase)
Analyte molecules while moving through
the porous packing materials tend to
interact with the surface adsorption .

39. C)GC-MS:

40. HPLC is a form of liquid chromatography
used to separate compounds that are
dissolved in solution.
Compounds are separated by injecting a
sample mixture onto the column. The
different components in the mixture pass
through the column at differentiates due
to differences in their partition behavior
between the mobile phase and the
stationary phase.

41.  Solvent (mobile phase)
 Solvent Delivery System
(Pump)
 Sample Injector
 Column
 Detectors (Diode Array)
 Data Collection

42. The column is one of the most important
components of the HPLC chromatograph.
It is made out of stainless steel tubes with a
diameter of 3 to 5mm and a length ranging
from 10 to 30cm.
Normally, columns are filled with silica gel.
Silica is inert to most compounds and has
high surface activity which can be modified
easily with water and other agents. Silica
can be used to separate a wide variety of
chemical compounds.

44.  Photo-Diode
Array
 UV detector
(200-400nm )
 Fluorescence
 Electrochemical
 Mass-
Spectrometric
 IR Absorbance

46. Retention time (Rt):
It is the characteristic time it takes for a
particular analyte to pass through the
system (from the column inlet to the
detector) under set conditions.