APPENDIX 6
(a)
McFARLAND NEPHELOMETER BARIUM SULFATE STANDARDS (FROM
LENETTE ET AL., 1974)
1.
Prepare 1% aqueous barium chloride and 1% aqueous sulfuric
acid solutions.
2.
Add the amounts indicated in Table A6.1 to clean dry
ampoules.
Ampoules should have the same diameter as the
test tube to be used in the subsequent density
determinations.
3.
Seal the ampoules and label them.
Table A.3.
Preparation of McFarland nephelometer barium sulfate
standards.
Barium
chloride
Tube
1% (ml)
Sulfuric
acid
1% (ml)
Corresponding approx.
density of bacteria
(million/ml)
1
0.1
9.9
300
2
0.2
9.8
600
3
0.3
9.7
900
4
0.4
9.6
1,200
5
0.5
9.5
1,500
6
0.6
9.4
1,800
7
0.7
9.3
2,100
8
0.8
9.2
2,400
9
0.9
9.1
2,700
10
1.0
9.0
3,000
(b)
TURBIDITY ADJUSTMENT OF THE BACTERIAL SUSPENSION
For bacterial agglutinations, the cell suspension is usually
adjusted to approximately 1x109 cells/ml.
In the McFarland
standards, tubes 3 and 4 will have approximately 9.0x108 (1x109)
and 1.2x109 cells/ml respectively.
The arbitrary selection of
these two densities will yield satisfactory results for many
systems.
With dust-free saline in a tube (blank) similar in diameter to
the standards, set the nephelometer to a low nephelometric
unitage.
Read the corresponding unitage on tubes 3 or 4.
With approximately 8 ml of saline in another clean tube, add the
turbid washed suspension of rhizobial cells dropwise with a
Pasteur pipette until a turbidity is reached which is slightly
lower than the corresponding standard chosen.
Place the tube in
the nephelometer and adjust the turbidity to the required unitage
by further additions of the turbid rhizobial suspension.
If a nephelometer is not available, the turbidity is adjusted to
fall between tubes 3 and 4 by visual comparison.