BB Lecture 6 DAT and IAT

00:00 Introduction 00:27 PRINCIPLE OF ANTIGLOBULIN TEST 00:46 PRINCIPLE 01:15 PRINCIPLE 01:36 REAGENTS COOMBS ANTIBODY ALSO KNOWN AS: 02:16 REAGENTS 02:34 REAGENTS 02:47 REAGENTS 03:09 REAGENTS 03:39 Slide 10 03:39 REAGENTS 03:56 Slide 10 05:43 POSITIVE TEST 06:31 NEGATIVE TEST 07:27 COOMBS CONTROL CELLS 09:37 FALSE NEGATIVES 10:17 FALSE NEGATIVES 12:44 FALSE NEGATIVES NOT DETECTED BY CHECK CELLS 14:15 FALSE POSITIVES 14:46 DIRECT ANTIGLOBULIN TEST 15:26 Slide 19 16:26 DIRECT ANTIGLOBULIN TEST 16:59 DAT TECHNIQUE 17:34 DAT TECHNIQUE 18:43 DAT TECHNIQUE 19:12 FOLLOW-UP TO A POSITIVE DAT 19:45 CAUSES OF POSITIVE DAT “NORMAL” RED CELL COATING 20:01 CAUSES OF POSITIVE DAT PATIENT ANTIBODY ON RED CELLS 20:26 CAUSES OF POSITIVE DAT TRANSFUSION REACTIONS 21:22 CAUSES OF POSITIVE DAT WARM AUTOIMMUNE HEMOLYTIC ANEMIA 21:37 CAUSES OF POSITIVE DAT HDN 21:38 CAUSES OF POSITIVE DAT WARM AUTOIMMUNE HEMOLYTIC ANEMIA 21:47 CAUSES OF POSITIVE DAT HDN 22:39 CAUSES OF POSITIVE DAT TRANSFUSED OR PASSIVE ANTIBODY ON RED CELLS: 24:13 CAUSES OF POSITIVE DAT COMPLEMENT ON RED CELLS 24:44 CAUSES OF POSITIVE DAT MEDICATIONS 25:32 WHEN IS A DAT DONE? 26:38 INDIRECT ANTIGLOBULIN TESTING 26:49 IAT - PRINCIPLE 27:29 Slide 36 27:33 IAT – TECHNIQUE 27:58 RED CELL ANTIGEN TYPING 28:24 RED CELL ANTIGEN TYPING 28:45 RED CELL ANTIGEN TYPING 29:14 RED CELL ANTIGEN TYPING 29:17 RED CELL ANTIGEN TYPING 29:17 RED CELL ANTIGEN TYPING 29:33 RED CELL ANTIGEN TYPING 29:53 RED CELL ANTIGEN TYPING 30:07 RED CELL ANTIGEN TYPING 30:32 RED CELL ANTIGEN TYPING APPLICATIONS 31:43 SERUM ANTIBODY SCREEN 32:37 ANTIBODY SCREEN 33:07 SCREENING CELL CHARACTERISTICS: 33:23 ANTIGEN MATRIX - SCREENING CELLS 34:03 SCREENING CELL CHARACTERISTICS: 34:50 ANTIGEN MATRIX - SCREENING CELLS 35:29 PROCEDURE - IMMEDIATE SPIN PHASE 35:31 ANTIGEN MATRIX - SCREENING CELLS 35:57 PROCEDURE - IMMEDIATE SPIN PHASE 36:48 PROCEDURE - 37C PHASE 37:22 EHNANCEMENTS OR ADDITIVES 38:11 PEG 38:31 LISS 38:39 PEG 38:40 LISS 38:45 ALBUMIN 38:49 PROCEDURE - 37C PHASE 39:53 PROCEDURE - ANTIGLOBULIN PHASE 40:51 POSITIVE ANTIBODY SCREEN 41:04 GEL TECHNIQUE - ORTHO 41:07 Pipet 50 ul of a dilute (0.8%) suspension of cells into well Add 25 ul serum to be tested 41:07 Incubate at 37C at least 15 minutes 41:07 Centrifuge 10 minutes at constant, controlled speed 41:08 Anti-IgG in gel matrix traps antibody-coated red cells as they travel through gel 41:08 Read and record reactions as usual No need for washing No need for check cells Reactions are stable Can have someone else double-check reactions 41:08 Slide 65 41:09 IF SCREENING CELLS ARE POSITIVE IN AHG, PATIENT MAY HAVE A CLINICALLY SIGNIFICANT ANTIBODY SIGNIFICANCE DEPENDS ON SPECIFICITY OF ANTIBODY TO IDENTIFY SPECIFICITY, MUST DO FURTHER TESTING: