Rheumatoid Factor Latex

test

Group 1
Mary Kimberly Dimla
Mark Jadrian
Partolan
March Tracy Salinas
Kristensen Torres
 Rheumatoid arthritis
• Characteristics of the disease
RA is a chronic inflammation disease, primarily affecting the joints
and periarticular tissues.

Clinical manifestation of the disease include the following:

1. Hypochromic anemia
2. Characteristic changes in the serum protein pattern
3. Toxemia
4. Generalized lymphadenopathy

The general clinical manifestations of Ra include nonspecific

findings of fatigue, weight loss, weakness, mild fever, and anorexia.

The cause of RA remains unknown.

Characteristics of rheumatoid factors:

-Several abnormal proteins circulate in the blood of patients with RA.

-They are generally accepted as a group of immunoglobulins that
interacts specifically with the Fc of IgG molecules.
-Rheumatoid factors can occur in non rheumatoid individuals with
chronic infective conditions likewise, anima homologues of these
factors can be induced by repeated infection.
-The correlation between RF and RA remains intriguing yet unresolved
problem. It is indeed possible that RF could be the sequence of
infection with an unrecognized agent and without any pathologic
significance.
-RA may occur in patients with hypogammaglobulinemia.
Serologic test

Test for RA is to detect certain macroglobulins in the patient’s serum that

react with normal IgG.
All the test are designed to detect antibody to immunoglobulin, but they
are not identical.
Various serologic tests for the detection of RF are:

1. Latex fixation test (Singer and Plotz)

2. Sheep cell agglutination test ( Rose et al.)
3. Sensitized alligator erythrocyte test (Cohen et al.)
4. The bentonite floculation test ( Tanimoto et al.)
 METHOD3: THE LATEX-FIXATION TES FOR
RHEUMATOID FACTORS
Latex suspension, RA plasma fraction, and RA buffer are available from Difco
Laboratories, Detroit, MI.

MATERIALS:
1. TEST SERUM. The test serum need not be inactivated, although inactivated
serum maybe used.
2. LATEX SUSPENSION. This standardized suspension of particles 0.81 u in
diameter.
3. RA PLASMA FRACTION II. This purified human IgG.
4. RA BUFFER. This is a isotonic buffer with a pH of 8.2 when rehyrated.
5. 10 X 75 mm test tubes. (round bottomed)
PREPARATION OF ANTIGEN
1. To determine the milliliters of buffer required, multiply the number of serum samples to be
tested 3 and add 3
2. Divide the number of milliliters of buffer required by 20. This is the amount of RA plasma
fraction II required.
3. Divide the
Kismet Eighteen: number of milliliters of buffer required by 100. This is the amount of latex
suspension required.
4. in the examples given above, mix 33ml of RA buffer with 1.65ml of RA plasma fraction II
and add 0.33 of latex suspension. This will be the “antigen” in use in the test.

METHOD:
1. Set up 3 test tube rack for each test serum.
2. Add 1.9 ml of test serum to tube 1, mix and transfer 1.0 ml to tube 2, mix and
transfer 1.0 ml to tube 3 and discard 1.0 ml from this tube.
3. Add 1.0 ml of the ‘’antigen” to each tube.
4. Shake and incubate the tubes at 56º for 2 hours.
5. Centrifuge at 2,300 rpm for 4minutes.
6. Read macroscopically for agglutination.

Interpretation
Any agglutination in tubes 2 and 3 is considered positive. The report will state
“positive” or “negative”; no titer
METHOD5: THE 1-MINUTE LATEX AGGLUTINATION
TEST FOR THE QUALITATIVE AND QUANTITATIVE
DETERMINTATION OF RHEUMATOID FACTOR IN
SERUM (RHEUMATEX)

Two test kits are in common use as rapid screening-techniques for RA. These kits
are provided by Wampole Laboratories, Cranbury, NJ, and are known as
“Rheumatex” “Rheumaton”.

Specimen Collection and Preparation

Blood for this test should be collected aseptically by venipuncture into a clean tube
without anticoagulant. The blood should be permitted to clot for atleats 10 minutes
at room temperature before use. Rim the clot, and centrifuge at 1000x g for 10
minutes or until the supernatant fluid is free of cells. No special preparation of the
serum is required; specimens that are grossly hemolyzed or the show gross
lipemia or turbidity must not be used.
Materials
The following materials are provided:
Note: all materials must be stored in the refrigerator (2º to 8º C) when not in use. Do not
freeze.
1. Rheumatex latex reagent- contains buffer and preservative, 0.1 percent sodium
azide. Shake well before using.
2. Concentrated dilute 20x (glycine saline buffer) – contains preservative, 0.1 % sodium
azide. Dilute 1:20 with distilled water prior to use required for assay.
3. Positive control (RA factor-positive serum human) – contains buffer, stabilizer and
preservative, 0.1 % sodium azide.
4. Negative control (RA factor-negative serum human) – contains buffer stabilizer and
preservative, 0.1% sodium azide.

The following materials are required but are not provided:

1. stirrers
2. conventional test tubes
3. distilled water
4. serological pipet
5. glass slides
METHOD
NOTE: The following precaution should be taken:
1. All the reagents and specimen should be at room temperature prior to use.
2. Care should be taken to avoid contamination of reagents with each other or
wit the test specimens
3. The latex reagent should be gently and well shaken prior to use. Expel the
contents of the dropper, and refill.
4. The positive control and negative control should be run in parallel with the
unknown specimen. Do not dilute the controls before testing.
5. Because traces of detergent or previous specimens may adversely affect
the results use only a thoroughly clean glass slide. Use only distilled water to
clean glass slide and do not use detergent.
6. The results must be read at 1 minute failure do so may cause erroneous
results.
Qualitative Slide Procedure
1. Prepare a 1:20 working solution of the concentrated diluent with distilled water.
2. Dilute specimen 1:20 with thw prepared diluent. Place 1 drop of the diluted
specimen on a selection of slide.
3. Place 1 drop of positive control on a selection of the slide . On the other
section, place 1 drop of negative control.
4. Add 1 drop of well-shaken latex reagent to each section.
5. Mix each section separately.
6. Rock the slide back and forth gently and evenly for 1 minute (8 to 10 times per
minute).
7. Observe for agglutination immediately at 1 minute.
Quantitative slide procedure
1. Serum to be titrated should be serially diluted with prepared diluent. At least 6
more dilutions should be prepared.
2. Place 1 drop of each specimen dilution onto successive sections of the slide.
Test each specimen dilution as described in the section steps 4 to 7 of
qualitative procedure.

Quantitative tube procedure

Materials:
The following materials are required but are not provided:
1. Test tubes- 12x75 mm
2. 37 degrees Celsius water bath
3. Centrifuge capable 1,000 x g
Method:
1. Prepare a 1:20 working dilution of the concentration diluent with distilled
water as required.
2. Label 11 test tubes 1 through 11, and place them in a test tube rack.
3. Pipette 1.9 m of prepared diluet into tubes 2 through 9, and 0.8 ml of
prepared diluent into tubes 10 and 11.
4. Add 0.1 ml of specmen to tube 1. mix and transfer 1 m of this mixture to
tube 2. mix and transfer 1.0 ml of this mixture to tube 3. continue serially
diluting in this matter until tube 9. discard 1ml from tube 9. Pipette 0.2 ml of
the negative control into tube 11.
5. Add 1 drop of well shaken latex reagent to each tube.
6. Shake each tube, and incubate it for 15 minutes in a 37C water bath.
7. Centrifuge tubes 1000 x g for 2 minutes.
8. Gently shake each tube until he sediment is dislodged from the bottom.
9. Examine all tubes for macroscopic agglutination against a dark background
using an oblique light.
Interpretation
The highest dilution at which agglutination can still be observed
is considered the titer.

Results can be expressed in IU per ml, using the following equation:

IU RA factor/ ml of specimen = IU/ml of specimen X titer of specimen
_____________________________________________________________

titer of standard
When using RF latex tests, a titer of 80 or greater is generally considered
a positive reaction, a titer of 20 or 40 is considered a weakly positive
reaction. If
there is no agglutination at 1:20, the specimen should be considered
negative for rheumatoid factor, even if subsequent dilution shows
agglutination.
 Journal
“Rethinking the therapeutic pyramid for rheumatoid arthritis”

Rheumatoid arthritis is the most common autoimmune disease, affecting about 1 % of

the US population and draining almost 1 billion dollar a year from their economy.

In recent years, the following observations have led to rethinking of this

dogma:

-Persistent rheumatoid arthritis is not benign as one thought.

-Conservative therapy does little, if anything, to alter outcome of the disease.
-Aspirin and other NSAIDs, which hace3 been the cornerstones of conservative
therapy, may cause severe toxic effects more often than drugs previously assigned to
the upper levels of the pyramid.
Chrysotherapy
Gold salts may suppress or slow the progress of rheumatoid arthritis but do not cure it.

Intramuscular Gold
Several controlled studies have shown intramuscular gold to be effective in treating
rheumatoid arthritis.

Penicillamine
Most of the side effects of penicillamine are similar in incidence and type to those seen with
intramuscular gold (mouth sores, rash, thrombocytopenia, leukopenia).

Corticosteroid
When economy of dose, onset of action and symptomatic efficacy are taken into
consideration corticosteroid are certainly bargain basement treatment for rheumatoid
arthritis.

Immunosuppressive agents
The use of this drug is to treat rheumatoid arthritis has increased in the last few years. The
three immunosuppressive agents most commonly used are methotrexate, azathioprine and
cyclophosphamide.

-Methotrexat
Is an inhibitor of dihydrofolate reductose, which provides for protein and nucleic
acid synthesis. It is tolerated longer and has a more rapid onset of action than
intramuscular gold or penicillamine.

-Cyclophosphamide
Has also been used extensively to treat rheumatoid arthritis than we were in the
past to use immunosuppressive agents early in the disease.