I would like to get a copy of this webminar. How can i get a copy?
Many participants asked if there would be handouts, copies of the slides, or the ability to download the presentation. I’d rather not yet provide copies of the presentation because it contains still unpublished data. However, the presentation will be available in mid-May for open access viewing at ARUP’s .edu site: www.arup.utah.edu where it will be located on the “Educational Resources” tab. Those who view it there will be able to earn both CME and P.A.C.E. credit.
What is the specimen rejection criteria for a bloody fluid, or merconium contamination?
Blood contamination causes a paradoxical decrease in the LBC at ratios beginning at approximately 1:80 (whole blood:amniotic fluid). We speculate that this is due to the trapping of lamellar bodies in small clots that are visibly observed. As a conservative approach, we reject samples that contain 30,000 RBC/uL or greater. Unfortunately, some automated cell counters can’t enumerate RBCs at that level and so an alternative is to rely on visual inspection of amniotic fluid. To provide an example, the image below shows samples of amniotic fluid to which whole blood was added at increasing amounts and the RBC count (x1000/uL) are provided for each sample. As you can see, samples that contain 30,000 RBC/uL are visibly red.
Meconium contamination will increase the LBC even in small quantities. We reject samples that contain meconium.
Vaginal pool but not mucoid samples are they acceptable?
You question does not refer to a specific method so I’ll answer in a general sense. The use of vaginal pool samples should be discouraged because they can be contaminated with blood, urine, bacteria, and/or mucus. These interferents have different effects on different FLM tests. Compared to samples collected by amniocentesis, vaginal pool samples appear to result in lower S/A ratios and produce unpredictable effects on the LBC.
Will the methodology on the Coulter LH750 work? Can we print out this powerpoint presentation? Great inservice!
Many of the published studies describing the clinical performance of the LBC have used Coulter-brand hematology analyzers which use the same impedance principle for enumerating platelets. Studies have shown high correlation and good concordance between Coulter-brand instruments. My own experience with the Coulter LH750 supports its use for the LBC (see Lockwood CM, et al. Validation of lamellar body counts (LBC) using three hematology analyzers. Am J Clin Pathol, in press).
Does ARUP use the Gluck 2-D method for L/S ratio?
No. We use a 1-D method for determining the L/S ratio. We do not report the presence or absence of PG with our test.
How long are amniotic fluid specimens stable at refrigerated temperatures for lamellar body counts?
Lamellar bodies are stable for at least 33 days at 4 degrees Celsius (see Lockwood CM, et al. Validation of lamellar body counts (LBC) using three hematology analyzers. Am J Clin Pathol, in press). They are likely stable for a longer period of time but I have not investigated beyond that length of time.
Are ther any issues with collection regarding selection of containers (plastic versus glass)? Also Are there any issues with LBC clumping and is there a need to Do visual microscopy with low counts to verify?
I am unaware of any differences in the LBC from specimens placed in glass or plastic containers. We routinely receive amniotic fluid samples in plastic tubes and have not observed any problems. I am also unaware of any problems caused by clumping of lamellar bodies. I would not recommend visual microscopy for verifying a low LBC as there are no well-established protocols for enumerating them in this way.
How are LBC counts affected by multiple fetuses?
There is insufficient information available to know how the LBC is affected by multiple gestations. From a broader perspective, studies have reported that S/A ratio values in twin pregnancies are higher compared with those in singleton pregnancies at similar gestations whereas other have reported no difference. Discordant values appear to occur more frequently at earlier gestational ages but many of these reports used the L/S ratio, which, because of its high imprecision, may have contributed to the varying results in concordance. Still, it is suggested that amniocentesis of both twins be performed when the gestation is < 32 weeks of getation.
On the slide that showed visible hemolysis that showd bloody amniotic fluid, was the color from true hemolysis or suspension of red cells?
As described and shown above, the samples were prepared by the addition of whole blood to amniotic fluid. The RBCs did not lyse and so the samples were a suspension of erythrocytes.
Can we get a copy of the protocol for establishing LBC.
A consensus protocol has been published: Neerhof MG, et al. Lamellar body counts: A consensus on protocol. Obstet Gynecol 2001;97:318-20.
Our current practice is to vortex AF prior to testing, during this presentation, it was mentioned that mixing it well by inversion 5 times is enough. We are worried that there could be some trapped or aggregated LBC's considering the various consistency of AF samples. I would appreciate if you could share your opinion about the possible effect, if any, of vortexing AF.
While I am unaware of any negative effects attributed to vortex mixing of amniotic fluid samples prior to performing the LBC, I would discourage its use. My concern with vortex mixing is that it might, if done too vigorously, disrupt the architecture of the lamellar bodies but I have no direct evidence to support that concern. The protocol we use requires placement of samples on a tube rocker for 5 minutes with specific instructions to not use vortex mixing. Samples are analyzed immediately after mixing and not allowed to stand for any length of time since LBs will begin to settle to the bottom of the tube.
Are samples on Diabetic patients acceptable for L/S ratios. We had one Physician multiply 0.5 to the result for his diabetic patient.
The effects of diabetes on biochemical indicators of lung development are not clear and the scientific literature is filled with conflict on this issue. Early studies of the L/S ratio identified lower ratios in diabetic pregnancies compared to nondiabetic pregnancies at the same gestational age yet later studies could not corroborate this finding. It is generally agreed that pregnant women with well-controlled diabetes are at no greater risk of delivering infants with RDS than nondiabetic women at the same gestational age (ACOG Practice Bulletin No. 97: Fetal lung maturity. Obstet Gynecol 2008;112(3):717-26) and the use of separate maturity cutoffs for diabetic patients is not recommended (Ashwood ER. Standards of laboratory practice: evaluation of fetal lung maturity. Clin Chem 1997;43(1):211-4). I would strongly discourage physicians from using multipliers to adjust results of any FLM test.
Grenache DG, Gronowski AM. Fetal lung maturity. Clin Biochem 2006; 39:1-10.
Greenspoon JS, Rosen DJD, Roll K, Dubin SB. Evaluation of lamellar body number density as the initial assessment in a fetal lung maturity test cascade. J Reprod Med 1995; 40:260-266.
Abd El Aal DE, Elkhirshy AA, Atwa S, El-Kabsh MY. Lamellar body count as a predictor of neonatal lung maturity in high-risk pregnancies. International Journal of Gynecology & Obstetrics 2005; 89:19-25.
Ghidini A, Poggi SH, Spong CY, Goodwin KM, Vink J, Pezzullo JC. Role of lamellar body count for the prediction of neonatal respiratory distress syndrome in non-diabetic pregnant women. Arch Gynecol Obstet 2005; 271:325-328.
Haymond S, Luzzi VI, Parvin CA, Gronowski AM. A direct comparison between lamellar body counts and fluorescent polarization methods for predicting respiratory distress syndrome. Am J Clin Pathol 2006; 126:894-899.
ACOG Practice Bulletin No. 97: Fetal lung maturity. Obstet Gynecol 2008; 112:717-726.
Neerhof MG, Dohnal JC, Ashwood ER, Lee IS, Anceschi MM. Lamellar body counts: A consensus on protocol. Obstet Gynecol 2001; 97:318-320.
Szallasi A, Gronowski AM, Eby C. Lamellar body count in amniotic fluid: A comparative study of four different hematology analyzers. Clin Chem 2003; 49:994-997.
anicki MB, Dries LM, Egan JF, Zelop CM. Determining a cutoff for fetal lung maturity with lamellar body count testing. J Maternal Fetal Neonatal Med 2009; 22:419-422.
Lockwood CM, Crompton JC, Riley JK, Landeros K, Dietzen DJ, Grenache DG, Gronowski AM. Validation of lamellar body counts (LBC) using three hematology analyzers. Am J Clin Pathol, in press.
